BACKGROUND: The Human Leukocyte Antigen (HLA) genes are a group of highly polymorphic genes that are located in the Major Histocompatibility Complex (MHC) region on chromosome 6. The HLA genotype affects the presentability of tumour antigens to the immune system. While knowledge of these genotypes is of utmost importance to study differences in immune responses between cancer patients, gold standard, PCR-derived genotypes are rarely available in large Next Generation Sequencing (NGS) datasets. Therefore, a variety of methods for in silico NGS-based HLA genotyping have been developed, bypassing the need to determine these genotypes with separate experiments. However, there is currently no consensus on the best performing tool. RESULTS: We evaluated 13 MHC class I and/or class II HLA callers that are currently available for free academic use and run on either Whole Exome Sequencing (WES) or RNA sequencing data. Computational resource requirements were highly variable between these tools. Three orthogonal approaches were used to evaluate the accuracy on several large publicly available datasets: a direct benchmark using PCR-derived gold standard HLA calls, a correlation analysis with population-based allele frequencies and an analysis of the concordance between the different tools. The highest MHC-I calling accuracies were found for Optitype (98.0%) and arcasHLA (99.4%) on WES and RNA sequencing data respectively, while for MHC-II HLA-HD was the most accurate tool for both data types (96.2% and 99.4% on WES and RNA data respectively). CONCLUSION: The optimal strategy for HLA genotyping from NGS data depends on the availability of either WES or RNA data, the size of the dataset and the available computational resources. If sufficient resources are available, we recommend Optitype and HLA-HD for MHC-I and MHC-II genotype calling respectively.

BACKGROUND: Genetic barcoding provides a high-throughput way to simultaneously track the frequencies of large numbers of competing and evolving microbial lineages. However making inferences about the nature of the evolution that is taking place remains a difficult task. RESULTS: Here we describe an algorithm for the inference of fitness effects and establishment times of beneficial mutations from barcode sequencing data, which builds upon a Bayesian inference method by enforcing self-consistency between the population mean fitness and the individual effects of mutations within lineages. By testing our inference method on a simulation of 40,000 barcoded lineages evolving in serial batch culture, we find that this new method outperforms its predecessor, identifying more adaptive mutations and more accurately inferring their mutational parameters. CONCLUSION: Our new algorithm is particularly suited to inference of mutational parameters when read depth is low. We have made Python code for our serial dilution evolution simulations, as well as both the old and new inference methods, available on GitHub ( https://github.com/FangfeiLi05/FitMut2 ), in the hope that it can find broader use by the microbial evolution community.

BACKGROUND: In many organisms increased reproductive effort is associated with a shortened life span. This trade-off is reflected in conserved molecular pathways that link nutrient-sensing with fecundity and longevity. Social insect queens apparently defy the fecundity / longevity trade-off as they are both, extremely long-lived and highly fecund. Here, we have examined the effects of a protein-enriched diet on these life-history traits and on tissue-specific gene expression in a termite species of low social complexity. RESULTS: On a colony level, we did not observe reduced lifespan and increased fecundity, effects typically seen in solitary model organisms, after protein enrichment. Instead, on the individual level mortality was reduced in queens that consumed more of the protein-enriched diet - and partially also in workers - while fecundity seemed unaffected. Our transcriptome analyses supported our life-history results. Consistent with life span extension, the expression of IIS (insulin/insulin-like growth factor 1 signalling) components was reduced in fat bodies after protein enrichment. Interestingly, however, genes involved in reproductive physiology (e.g., vitellogenin) were largely unaffected in fat body and head transcriptomes. CONCLUSION: These results suggest that IIS is decoupled from downstream fecundity-associated pathways, which can contribute to the remoulding of the fecundity/longevity trade-off in termites as compared to solitary insects.

BACKGROUND: Lineage distribution of Mycobacterium tuberculosis (Mtb) isolates is strongly associated with geographically distinct human populations, and its transmission can be further impacted by the bacterial genome. However, the epidemic success of Mtb isolates at an individual level was unknown in eastern China. Knowledge regarding the emergence and transmission of Mtb isolates as well as relevant factors may offer a new solution to curb the spread of the disease. Thus, this study aims to reveal the evolution and epidemic success of Mtb isolates in eastern China. RESULTS: Of initial 1040 isolates, 997 were retained after removing duplicates and those with insufficient sequencing depth. Of the final samples, 733 (73.52%) were from Zhejiang Province, and 264 (26.48%) were from Shanghai City. Lineage 2 and lineage 4 accounted for 80.44% and 19.56%, with common ancestors dating around 7017 years ago and 6882 years ago, respectively. Sub-lineage L2.2 (80.34%) contributed the majority of total isolates, followed by L4.4 (8.93%) and L4.5 (8.43%). Additionally, 51 (5.12%) isolates were identified to be multidrug-resistant (MDR), of which 21 (29.17%) were pre-extensively drug-resistant (pre-XDR). One clade harboring katG S315T mutation may date back to 65 years ago and subsequently acquired mutations conferring resistance to another five antibiotic drugs. The prevalence of compensatory mutation was the highest in pre-XDR isolates (76.19%), followed by MDR isolates (47.06%) and other drug-resistant isolates (20.60%). Time-scaled haplotypic density analyses suggested comparable success indices between lineage 2 and lineage 4 (P = 0.306), and drug resistance did not significantly promote the transmission of Mtb isolates (P = 0.340). But for pre-XDR isolates, we found a higher success index in those with compensatory mutations (P = 0.025). Mutations under positive selection were found in genes associated with resistance to second-line injectables (whiB6) and drug tolerance (prpR) in both lineage 2 and lineage 4. CONCLUSIONS: Our study demonstrates the population expansion of lineage 2 and lineage 4 in eastern China, with comparable transmission capacity, while accumulation of resistance mutations does not necessarily facilitate the success of Mtb isolates. Compensatory mutations usually accompany drug resistance and significantly contribute to the epidemiological transmission of pre-XDR strains. Prospective molecular surveillance is required to further monitor the emergence and spread of pre-XDR/XDR strains in eastern China.

BACKGROUND: Semen cryopreservation is a critical tool for breed improvement and preservation of biodiversity. However, instability of sperm freezability affects its application. The Mediterranean buffalo is one of the river-type buffaloes with the capacity for high milk production. Until now, there is no specific cryopreservation system for Mediterranean buffalo, which influences the promotion of excellent cultivars. To improve the semen freezing extender used in cryopreservation of Mediterranean buffalo, different protein datasets relating to freezability sperm were analyzed by iTRAQ-based proteomics. This study will be beneficial for further understanding the sperm freezability mechanism and developing new cryopreservation strategy for buffalo semen. RESULTS: 2652 quantified proteins were identified, including 248 significantly differentially expressed proteins (DEP). Gene Ontology (GO) analysis indicated that many these were mitochondrial proteins, enriched in the molecular function of phospholipase A2 activity and enzyme binding, and biological processes of regulation of protein kinase A signaling and motile cilium assembly. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis identified 17 significant pathways, including oxidative phosphorylation (OXPHOS). Furthermore, 7 DEPs were verified using parallel reaction monitoring or western blot, which confirmed the accuracy of the iTRAQ data. Peroxiredoxin 6 (PRDX6), which expressed 1.72-fold higher in good freezability ejaculate (GFE) compared to poor freezability ejaculate (PFE) sperms, was selected to explore the function in sperm freezability by adding recombinant PRDX6 protein into the semen freezing extender. The results showed that the motility, mitochondrial function and in vitro fertilization capacity of frozen-thawed sperm were significantly increased, while the oxidation level was significantly decreased when 0.1 mg/L PRDX6 was added compared with blank control. CONCLUSIONS: Above results revealed the metabolic pattern of freezability of Mediterranean buffalo sperms was negatively associated with OXPHOS, and PRDX6 had protective effect on cryo-damage of frozen-thawed sperms.

BACKGROUND: Sex determination is the process whereby the bipotential embryonic gonads become committed to differentiate into testes or ovaries. In genetic sex determination (GSD), the sex determining trigger is encoded by a gene on the sex chromosomes, which activates a network of downstream genes; in mammals these include SOX9, AMH and DMRT1 in the male pathway, and FOXL2 in the female pathway. Although mammalian and avian GSD systems have been well studied, few data are available for reptilian GSD systems. RESULTS: We conducted an unbiased transcriptome-wide analysis of gonad development throughout differentiation in central bearded dragon (Pogona vitticeps) embryos with GSD. We found that sex differentiation of transcriptomic profiles occurs at a very early stage, before the gonad consolidates as a body distinct from the gonad-kidney complex. The male pathway genes dmrt1 and amh and the female pathway gene foxl2 play a key role in early sex differentiation in P. vitticeps, but the central player of the mammalian male trajectory, sox9, is not differentially expressed in P. vitticeps at the bipotential stage. The most striking difference from GSD systems of other amniotes is the high expression of the male pathway genes amh and sox9 in female gonads during development. We propose that a default male trajectory progresses if not repressed by a W-linked dominant gene that tips the balance of gene expression towards the female trajectory. Further, weighted gene expression correlation network analysis revealed novel candidates for male and female sex differentiation. CONCLUSION: Our data reveal that interpretation of putative mechanisms of GSD in reptiles cannot solely depend on lessons drawn from mammals.

BACKGROUND: Alternative splicing is an important step in gene expression, generating multiple isoforms for the same genes and greatly expanding the diversity of proteomes. Genetic variation in alternative splicing contributes to phenotypic diversity in natural populations. However, the genetic basis of variation in alternative splicing in livestock including pigs remains poorly understood. RESULTS: In this study, using a Duroc x Pietrain F2 pig population, we performed genome-wide analysis of alternative splicing estimated from stranded RNA-Seq data in skeletal muscle. We characterized the genetic architecture of alternative splicing and compared its basic features with those of overall gene expression. We detected a large number of novel alternative splicing events that were not previously annotated. We found heritability of quantitative alternative splicing scores (percent spliced in or PSI) to be lower than that of overall gene expression. In addition, heritabilities showed little correlation between alternative splicing and overall gene expression. We mapped expression QTLs (eQTLs) and splice QTLs (sQTLs) and found them to be largely non-overlapping. Finally, we integrated sQTL mapping with phenotype QTL (pQTL mapping to identify potential mediator of pQTL effect by alternative splicing. CONCLUSIONS: Our results suggest that regulatory variation exists at multiple levels and that their genetic controls are distinct, offering opportunities for genetic improvement.

BACKGROUND: Understanding the genetic mechanisms underlying coat color inheritance has always been intriguing irrespective of the animal species including American mink (Neogale vison). The study of color inheritance in American mink is imperative since fur color is a deterministic factor for the success of mink industry. However, there have been no studies during the past few decades using in-depth pedigree for analyzing the inheritance pattern of colors in American mink. METHODS: In this study, we analyzed the pedigree of 23,282 mink extending up to 16 generations. All animals that were raised at the Canadian Center for Fur Animal Research (CCFAR) from 2003 to 2021 were used in this study. We utilized the Mendelian ratio and Chi-square test to investigate the inheritance of Dark (9,100), Pastel (5,161), Demi (4,312), and Mahogany (3,358) colors in American mink. RESULTS: The Mendelian inheritance ratios of 1:1 and 3:1 indicated heterozygous allelic pairs responsible for all studied colors. Mating sire and dam of the same color resulted in the production of offspring with the same color most of the time. CONCLUSION: Overall, the results suggested that color inheritance was complex and subjected to a high degree of diversity in American mink as the genes responsible for all four colors were found to be heterozygous.

BACKGROUND: Microsporidia are diverse spore forming, fungal-related obligate intracellular pathogens infecting a wide range of hosts. This diversity is reflected at the genome level with sizes varying by an order of magnitude, ranging from less than 3 Mb in Encephalitozoon species (the smallest known in eukaryotes) to more than 50 Mb in Edhazardia spp. As a paradigm of genome reduction in eukaryotes, the small Encephalitozoon genomes have attracted much attention with investigations revealing gene dense, repeat- and intron-poor genomes characterized by a thorough pruning of molecular functions no longer relevant to their obligate intracellular lifestyle. However, because no Encephalitozoon genome has been sequenced from telomere-to-telomere and since no methylation data is available for these species, our understanding of their overall genetic and epigenetic architectures is incomplete. METHODS: In this study, we sequenced the complete genomes from telomere-to-telomere of three human-infecting Encephalitozoon spp. -E. intestinalis ATCC 50506, E. hellem ATCC 50604 and E. cuniculi ATCC 50602- using short and long read platforms and leveraged the data generated as part of the sequencing process to investigate the presence of epigenetic markers in these genomes. We also used a mixture of sequence- and structure-based computational approaches, including protein structure prediction, to help identify which Encephalitozoon proteins are involved in telomere maintenance, epigenetic regulation, and heterochromatin formation. RESULTS: The Encephalitozoon chromosomes were found capped by TTAGG 5-mer telomeric repeats followed by telomere associated repeat elements (TAREs) flanking hypermethylated ribosomal RNA (rRNA) gene loci featuring 5-methylcytosines (5mC) and 5-hemimethylcytosines (5hmC), themselves followed by lesser methylated subtelomeres and hypomethylated chromosome cores. Strong nucleotide biases were identified between the telomeres/subtelomeres and chromosome cores with significant changes in GC/AT, GT/AC and GA/CT contents. The presence of several genes coding for proteins essential to telomere maintenance, epigenetic regulation, and heterochromatin formation was further confirmed in the Encephalitozoon genomes. CONCLUSION: Altogether, our results strongly support the subtelomeres as sites of heterochromatin formation in Encephalitozoon genomes and further suggest that these species might shutdown their energy-consuming ribosomal machinery while dormant as spores by silencing of the rRNA genes using both 5mC/5hmC methylation and facultative heterochromatin formation at these loci.

BACKGROUND: Since the NHGRI-EBI Catalog of human genome-wide association studies was established by NHGRI in 2008, research on it has attracted more and more researchers as the amount of data has grown rapidly. Easy-to-use, open-source, general-purpose programs for accessing the NHGRI-EBI Catalog of human genome-wide association studies are in great demand for current Python data analysis pipeline. RESULTS: In this work we present pandasGWAS, a Python package that provides programmatic access to the NHGRI-EBI Catalog of human genome-wide association studies. Instead of downloading all data locally, pandasGWAS queries data based on input criteria and handles paginated data gracefully. The data is then transformed into multiple associated pandas.DataFrame objects according to its hierarchical relationships, which makes it easy to integrate into current Python-based data analysis toolkits. CONCLUSIONS: pandasGWAS is an open-source Python package that provides the first Python client interface to the GWAS Catalog REST API. Compared with existing tools, the data structure of pandasGWAS is more consistent with the design specification of GWAS Catalog REST API, and provides many easy-to-use mathematical symbol operations.

BACKGROUND: Plant sexual reproduction is highly sensitive to elevated ambient temperatures, impacting seed development and production. We previously phenotyped this effect on three rapeseed cultivars (DH12075, Topas DH4079, and Westar). This work describes the transcriptional response associated with the phenotypic changes induced by heat stress during early seed development in Brassica napus. RESULTS: We compared the differential transcriptional response in unfertilized ovules and seeds bearing embryos at 8-cell and globular developmental stages of the three cultivars exposed to high temperatures. We identified that all tissues and cultivars shared a common transcriptional response with the upregulation of genes linked to heat stress, protein folding and binding to heat shock proteins, and the downregulation of cell metabolism. The comparative analysis identified an enrichment for a response to reactive oxygen species (ROS) in the heat-tolerant cultivar Topas, correlating with the phenotypic changes. The highest heat-induced transcriptional response in Topas seeds was detected for genes encoding various peroxidases, temperature-induced lipocalin (TIL1), or protein SAG21/LEA5. On the contrary, the transcriptional response in the two heat-sensitive cultivars, DH12075 and Westar, was characterized by heat-induced cellular damages with the upregulation of genes involved in the photosynthesis and plant hormone signaling pathways. Particularly, the TIFY/JAZ genes involved in jasmonate signaling were induced by stress, specifically in ovules of heat-sensitive cultivars. Using a weighted gene co-expression network analysis (WGCNA), we identified key modules and hub genes involved in the heat stress response in studied tissues of either heat-tolerant or sensitive cultivars. CONCLUSIONS: Our transcriptional analysis complements a previous phenotyping analysis by characterizing the growth response to elevated temperatures during early seed development and reveals the molecular mechanisms underlying the phenotypic response. The results demonstrated that response to ROS, seed photosynthesis, and hormonal regulation might be the critical factors for stress tolerance in oilseed rape.

BACKGROUND: N6-methyladenosine (m6A) refers to the methylation modification of N6 position of RNA adenine, a dynamic reversible RNA epigenetic modification that plays an important regulatory role in a variety of life processes. In this study, we used MeRIP-Seq and RNA-Seq of the longissimus dorsi (LD) muscle of adult (QA) and newborn (QN) Queshan Black pigs to screen key genes with m6A modification involved in muscle growth by bioinformatics analysis. RESULTS: A total of 23,445 and 25,465 m6A peaks were found in the whole genomes of QA and QN, respectively. Among them, 613 methylation peaks were significantly different (DMPs) and 579 genes were defined as differentially methylated genes (DMGs). Compared with the QN group, there were 1,874 significantly differentially expressed genes (DEGs) in QA group, including 620 up-regulated and 1,254 down-regulated genes. In order to investigate the relationship between m6A and mRNA expression in the muscle of Queshan Black pigs at different periods, a combined analysis of MeRIP-Seq and RNA-Seq showed that 88 genes were significantly different at both levels. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes results showed that DEGs and DMGs were mainly involved in skeletal muscle tissue development, FoxO signaling pathway, MAPK signaling pathway, insulin signaling pathway, PI3K-Akt signaling pathway, and Wnt signaling pathway. Four DEGs (IGF1R, CCND2, MYOD1 and FOS) and four DMGs (CCND2, PHKB, BIN1 and FUT2), which are closely related to skeletal muscle development, were selected as candidate genes for verification, and the results were consistent with the sequencing results, which indicated the reliability of the sequencing results. CONCLUSIONS: These results lay the foundation for understanding the specific regulatory mechanisms of growth in Queshan Black pigs, and provide theoretical references for further research on the role of m6A in muscle development and breed optimization selection.

BACKGROUND: DNA hypermethylation at promoter CpG islands (CGIs) is a hallmark of cancers and could lead to dysregulation of gene expression in the development of cancers, however, its dynamics and regulatory mechanisms remain elusive. Bivalent genes, that direct development and differentiation of stem cells, are found to be frequent targets of hypermethylation in cancers. RESULTS: Here we performed comprehensive analysis across multiple cancer types and identified that the decrease in H3K4me1 levels coincides with DNA hypermethylation at the bivalent promoter CGIs during tumorigenesis. Removal of DNA hypermethylation leads to increment of H3K4me1 at promoter CGIs with preference for bivalent genes. Nevertheless, the alteration of H3K4me1 by overexpressing or knockout LSD1, the demethylase of H3K4, doesn't change the level or pattern of DNA methylation. Moreover, LSD1 was found to regulate the expression of a bivalent gene OVOL2 to promote tumorigenesis. Knockdown of OVOL2 in LSD1 knockout HCT116 cells restored the cancer cell phenotype. CONCLUSION: In summary, our work identified a universal indicator that can pre-mark DNA hypermethylation in cancer cells, and dissected the interplay between H3K4me1 and DNA hypermethylation in detail. Current study also reveals a novel mechanism underlying the oncogenic role of LSD1, providing clues for cancer therapies.

BACKGROUND: Our knowledge about the ecological role of bacterial antimicrobial peptides (bacteriocins) in the human gut is limited, particularly in relation to their role in the diversification of the gut microbiota during early life. The aim of this paper was therefore to address associations between bacteriocins and bacterial diversity in the human gut microbiota. To investigate this, we did an extensive screening of 2564 healthy human gut metagenomes for the presence of predicted bacteriocin-encoding genes, comparing bacteriocin gene presence to strain diversity and age. RESULTS: We found that the abundance of bacteriocin genes was significantly higher in infant-like metagenomes (< 2 years) compared to adult-like metagenomes (2-107 years). By comparing infant-like metagenomes with and without a given bacteriocin, we found that bacteriocin presence was associated with increased strain diversities. CONCLUSIONS: Our findings indicate that bacteriocins may play a role in the strain diversification during the infant gut microbiota establishment.

As one of the important traits in pig production, meat quality has important research significance and value. Intramuscular fat (IMF) content is one of the most important factors affecting pork quality. Many experimental studies have shown that IMF content is closely related to the flavor, tenderness, and juiciness of pork. Therefore, it is of great significance to study the mechanism of porcine IMF deposition. Previous research indicated that miR-149-5p promoted the proliferation of porcine intramuscular (IM) preadipocytes and decreased their ability to differentiate, albeit the exact mechanism of action is unknown. In vitro, foreign pigs showed increased miR-149-5p expression and reduced fat deposition when compared to Queshan Black pigs. This study conducted metabolomics and transcriptomics analyses of porcine IM preadipocytes overexpressing miR-149-5p to verify their effects on lipid formation. According to metabolomics analysis, the overexpression of miR-149-5p has significantly altered the lipid, organic acid, and organic oxygen metabolites of porcine IM preadipocytes. Specially speaking, it has changed 115 metabolites, including 105 up-regulated and 10 down-regulated ones, as well as the composition of lipid, organic acid, and organic oxygen metabolism-related metabolites. RNA-seq analysis showed that overexpression of miR-149-5p significantly altered 857 genes, of which 442 were up-regulated, and 415 were down-regulated, with enrichment to MAPK, IL-17, PI3K-Akt, and ErbB signaling pathways. We found that overexpression of miR-149-5p inhibited adipogenic differentiation by changing cAMP signaling pathway in porcine IM preadipocytes. In addition, the overexpression of miR-149-5p may affect the transport of Cu2+ by targeting ATP7A and inhibiting adipogenic differentiation. These findings elucidate the regulatory function of miR-149-5p in porcine IM preadipocytes, which may be a key target for controlling pork quality.

BACKGROUND: Asthma is a common chronic respiratory disease worldwide. Recent studies have revealed the critical effects of the ceRNA network and ferroptosis on patients with asthma. Thus, this study aimed to explore the potential ferroptosis-related ceRNA network, investigate the immune cell infiltration level in asthma through integrated analysis of public asthma microarray datasets, and find suitable diagnostic biomarkers for asthma. METHODS: First, three asthma-related datasets which were downloaded from the Gene Expression Omnibus (GEO) database were integrated into one pooled dataset after correcting for batch effects. Next, we screened differentially expressed lncRNAs (DElncRNAs) between patients and healthy subjects, constructed a ceRNA network using the StarBase database and screened ferroptosis-related genes from the predicted target mRNAs for Disease Ontology (DO), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. We also performed Gene Set Enrichment Analysis (GSEA) and Gene Set Variation Analysis (GSVA) on the batch effect-corrected mRNA expression profile. Then, Least Absolute Shrinkage and Selection Operator (LASSO) regression was used to screen potential diagnostic biomarkers, and the diagnostic efficacy was assessed using a receiver operating characteristic (ROC) curve. Finally, we determined the proportion of 22 immune cells in patients with asthma using CIBERSORT and investigated the correlation between key RNAs and immune cells. RESULTS: We obtained 19 DElncRNAs, of which only LUCAT1 and MIR222HG had corresponding target miRNAs. The differentially expressed ferroptosis-related genes were involved in multiple programmed cell death-related pathways. We also found that the mRNA expression profile was primarily enriched in innate immune system responses. We screened seven candidate diagnostic biomarkers for asthma using LASSO regression (namely, BCL10, CD300E, IER2, MMP13, OAF, TBC1D3, and TMEM151A), among which the area under the curve (AUC) value for CD300E and IER2 were 0.722 and 0.856, respectively. Finally, we revealed the infiltration ratio of different immune cells in asthma and found a correlation between LUCAT1, MIR222HG, CD300E, and IER2 with some immune cells. CONCLUSION: This study explored a potential lncRNA-miRNA-mRNA regulatory network and its underlying diagnostic biomarkers (CD300E and IER2) in asthma and identified the immune cells most associated with them, providing possible diagnostic markers and immunotherapeutic targets for asthma.

BACKGROUND: Hong Kong catfish (Clarias fuscus) is an ecologically and economically important species that is widely distributed in freshwater regions of southern China. Hong Kong catfish has significant sexual growth dimorphism. The genome assembly of the Hong Kong catfish would facilitate study of the sex determination and evolution mechanism of the species. RESULTS: The first high-quality chromosome-level genome of the Hong Kong catfish was constructed. The total genome was 933.4 Mb, with 416 contigs and a contig N50 length of 8.52 Mb. Using high-throughput chromosome conformation capture (Hi-C) data, the genome assembly was divided into 28 chromosomes with a scaffold N50 length of 36.68 Mb. A total of 23,345 protein-coding genes were predicted in the genome, and 94.28% of the genes were functionally annotated in public databases. Phylogenetic analysis indicated that C. fuscus and Clarias magur diverged approximately 63.7 million years ago. The comparative genome results showed that a total of 60 unique, 353 expanded and 851 contracted gene families were identified in Hong Kong catfish. A sex-linked quantitative trait locus identified in a previous study was located in a sex-determining region of 30.26 Mb (0.02 to 30.28 Mb) on chromosome 13 (Chr13), the predicted Y chromosome. This QTL region contained 785 genes, of which 18 were identified as sex-related genes. CONCLUSIONS: This study is the first to report the chromosome-level genome assembly of Hong Kong catfish. The study provides an excellent genetic resource that will facilitate future studies of sex determination mechanisms and evolution in fish.

BACKGROUND: In the evolutionary study of gene families, exploring the duplication mechanisms of gene families helps researchers understand their evolutionary history. The tubby-like protein (TLP) family is essential for growth and development in plants and animals. Much research has been done on its function; however, limited information is available with regard to the evolution of the TLP gene family. Herein, we systematically investigated the evolution of TLP genes in seven representative Poaceae lineages. RESULTS: Our research showed that the evolution of TLP genes was influenced not only by whole-genome duplication (WGD) and dispersed duplication (DSD) but also by transposed duplication (TRD), which has been neglected in previous research. For TLP family size, we found an evolutionary pattern of progressive shrinking in the grass family. Furthermore, the evolution of the TLP gene family was at least affected by evolutionary driving forces such as duplication, purifying selection, and base mutations. CONCLUSIONS: This study presents the first comprehensive evolutionary analysis of the TLP gene family in grasses. We demonstrated that the TLP gene family is also influenced by a transposed duplication mechanism. Several new insights into the evolution of the TLP gene family are presented. This work provides a good reference for studying gene evolution and the origin of duplication.

BACKGROUND: Feed restriction occurs frequently during pig growth, either due to economic reasons or stressful environmental conditions. Local breeds are suggested to have better tolerance to periods of feed restriction. However, the mechanisms underlying the response to feed restriction in different breeds is largely unknown. The aims of the present study were (1) to compare the blood transcriptome profile in response to feed restriction and refeeding of two contrasted breeds, Large White (LW), which has been selected for high performance, and Creole (CR), which is adapted to tropical conditions, and (2) to investigate the effect of a moderate feed restriction and refeeding on whole blood transcriptome. Analysis of blood transcriptome allows to study the response to feed restriction and refeeding in a dynamic way. RNAseq was performed on blood samples of growing LW and CR pigs at two time points: after 3 weeks of feed restriction and after 3 weeks of refeeding. The data was compared with samples from control animals offered the same diet on an ad libitum basis throughout the whole experiment. RESULTS: In terms of performance (body weight and feed efficiency), CR pigs were less impacted by feed restriction than LW. The transcriptional response to feed restriction and refeeding between CR and LW was contrasted both in terms of number of DEGs and enriched pathways. CR demonstrated a stronger transcriptional response to feed restriction whereas LW had a stronger response to refeeding. Differences in the transcriptional response to feed restriction between CR and LW were related to cell stress response (Aldosterone Signalling, Protein ubiquitination, Unfolded Protein Signalling) whereas after refeeding, differences were linked to thermogenesis, metabolic pathways and cell proliferation (p38 MAPK, ERK/MAPK pathway). In both breeds, transcriptional changes related to the immune response were found after restriction and refeeding. CONCLUSIONS: Altogether, the present study indicates that blood transcriptomics can be a useful tool to study differential genetic response to feed restriction in a dynamic way. The results indicate a differential response of blood gene expression to feed restriction and refeeding between breeds, affecting biological pathways that are in accordance with performance and thermoregulatory results.

BACKGROUND: Sorghum (Sorghum bicolor [L.] Moench) is a promising target for pro-vitamin A biofortification as it is a global staple crop, particularly in regions where vitamin A deficiency is prevalent. As with most cereal grains, carotenoid concentrations are low in sorghum, and breeding could be a feasible strategy to increase pro-vitamin A carotenoids to biologically relevant concentrations. However, there are knowledge gaps in the biosynthesis and regulation of sorghum grain carotenoids, which can limit breeding effectiveness. The aim of this research was to gain an understanding of the transcriptional regulation of a priori candidate genes in carotenoid precursor, biosynthesis, and degradation pathways. RESULTS: We used RNA sequencing of grain to compare the transcriptional profile of four sorghum accessions with contrasting carotenoid profiles through grain development. Most a priori candidate genes involved in the precursor MEP, carotenoid biosynthesis, and carotenoid degradation pathways were found to be differentially expressed between sorghum grain developmental stages. There was also differential expression of some of the a priori candidate genes between high and low carotenoid content groups at each developmental time point. Among these, we propose geranyl geranyl pyrophosphate synthase (GGPPS), phytoene synthase (PSY), and phytoene desaturase (PDS) as promising targets for pro-vitamin A carotenoid biofortification efforts in sorghum grain. CONCLUSIONS: A deeper understanding of the controls underlying biosynthesis and degradation of sorghum grain carotenoids is needed to advance biofortification efforts. This study provides the first insights into the regulation of sorghum grain carotenoid biosynthesis and degradation, suggesting potential gene targets to prioritize for molecular breeding.

BACKGROUND: The reduction in phenotypic performance of a population due to mating between close relatives is called inbreeding depression. The genetic background of inbreeding depression for semen traits is poorly understood. Thus, the objectives were to estimate the effect of inbreeding and to identify genomic regions underlying inbreeding depression of semen traits including ejaculate volume (EV), sperm concentration (SC), and sperm motility (SM). The dataset comprised ~ 330 K semen records from ~ 1.5 K Holstein bulls genotyped with 50 K single nucleotide polymorphism (SNP) BeadChip. Genomic inbreeding coefficients were estimated using runs of homozygosity (i.e., FROH > 1 Mb) and excess of SNP homozygosity (FSNP). The effect of inbreeding was estimated by regressing phenotypes of semen traits on inbreeding coefficients. Associated variants with inbreeding depression were also detected by regressing phenotypes on ROH state of the variants. RESULTS: Significant inbreeding depression was observed for SC and SM (p < 0.01). A 1% increase in FROH reduced SM and SC by 0.28% and 0.42% of the population mean, respectively. By splitting FROH into different lengths, we found significant reduction in SC and SM due to longer ROH, which is indicative of more recent inbreeding. A genome-wide association study revealed two signals positioned on BTA 8 associated with inbreeding depression of SC (p < 0.00001; FDR < 0.02). Three candidate genes of GALNTL6, HMGB2, and ADAM29, located in these regions, have established and conserved connections with reproduction and/or male fertility. Moreover, six genomic regions on BTA 3, 9, 21 and 28 were associated with SM (p < 0.0001; FDR < 0.08). These genomic regions contained genes including PRMT6, SCAPER, EDC3, and LIN28B with established connections to spermatogenesis or fertility. CONCLUSIONS: Inbreeding depression adversely affects SC and SM, with evidence that longer ROH, or more recent inbreeding, being especially detrimental. There are genomic regions associated with semen traits that seems to be especially sensitive to homozygosity, and evidence to support some from other studies. Breeding companies may wish to consider avoiding homozygosity in these regions for potential artificial insemination sires.

BACKGROUND: Scale insects are worldwide sap-sucking parasites, which can be distinguished into neococcoids and non-neococcoids. Neococcoids are monophyletic with a peculiar reproductive system, paternal genome elimination (PGE). Different with neococcoids, Iceryini, a tribe in non-neococcoids including several damaging pests, has abdominal spiracles, compound eyes in males, relatively abundant wax, unique hermaphrodite system, and specific symbionts. However, the current studies on the gene resources and genomic mechanism of scale insects are mainly limited in the neococcoids, and lacked of comparison in an evolution frame. RESULT: We sequenced and de novo assembled a transcriptome of Icerya aegyptiaca (Douglas), a worldwide pest of Iceryini, and used it as representative of non-neococcoids to compare with the genomes or transcriptomes of other six species from different families of neococcoids. We found that the genes under positive selection or negative selection intensification (simplified as "selected genes" below) in I. aegyptiaca included those related to neurogenesis and development, especially eye development. Some genes related to fatty acid biosynthesis were unique in its transcriptome with relatively high expression and not detected in neococcoids. These results may indicate a potential link to the unique structures and abundant wax of I. aegyptiaca compared with neococcoids. Meanwhile, genes related to DNA repair, mitosis, spindle, cytokinesis and oogenesis, were included in the selected genes in I. aegyptiaca, which is possibly associated with cell division and germ cell formation of the hermaphrodite system. Chromatin-related process were enriched from selected genes in neococcoids, along with some mitosis-related genes also detected, which may be related to their unique PGE system. Moreover, in neococcoid species, male-biased genes tend to undergo negative selection relaxation under the PGE system. We also found that the candidate horizontally transferred genes (HTGs) in the scale insects mainly derived from bacteria and fungi. bioD and bioB, the two biotin-synthesizing HTGs were exclusively found in the scale insects and neococcoids, respectively, which possibly show potential demand changes in the symbiotic relationships. CONCLUSION: Our study reports the first I. aegyptiaca transcriptome and provides preliminary insights for the genetic change of structures, reproductive systems and symbiont relationships at an evolutionary aspect. This will provide a basis for further research and control of scale insects.

BACKGROUND: The purpose of this study is to investigate the association of rotating night shift work, CLOCK, MTNR1A, MTNR1B genes polymorphisms and their interactions with type 2 diabetes among steelworkers. METHODS: A case-control study was conducted in the Tangsteel company in Tangshan, China. The sample sizes of the case group and control group were 251 and 451, respectively. The logistic regression, log-linear model and generalized multifactor dimensionality (GMDR) method were used to investigate the interaction between circadian clock gene, melatonin receptor genes and rotating night shift work on type 2 diabetes among steelworkers. Relative excess risk due to interaction (RERI) and attributable proportions (AP) were used to evaluate additive interactions. RESULTS: Rotating night shift work, current shift status, duration of night shifts, and average frequency of night shifts were associated with an increased risk of type 2 diabetes after adjustment for confounders. Rs1387153 variants in MTNR1B was found to be associated with an increased risk of type 2 diabetes, which was not found between MTNR1A gene rs2119882 locus, CLOCK gene rs1801260 locus and the risk of type 2 diabetes. The association between rotating night shift work and risk of type 2 diabetes appeared to be modified by MTNR1B gene rs1387153 locus (RERI = 0.98, (95% CI, 0.40-1.55); AP = 0.60, (95% CI, 0.07-1.12)). The interaction between MTNR1A gene rs2119882 locus and CLOCK gene rs1801260 locus was associated with the risk of type 2 diabetes (RERI = 1.07, (95% CI, 0.23-1.91); AP = 0.77, (95% CI, 0.36-1.17)). The complex interaction of the MTNR1A-MTNR1B-CLOCK-rotating night shift work model based on the GMDR methods may increase the risk of type 2 diabetes (P = 0.011). CONCLUSIONS: Rotating night shift work and rs1387153 variants in MTNR1B were associated with an increased risk of type 2 diabetes among steelworkers. The complex interaction of MTNR1A-MTNR1B-CLOCK-rotating night shift work may increase the risk of type 2 diabetes.

BACKGROUND: Chlamydia (C.) psittaci, the causative agent of avian chlamydiosis and human psittacosis, is a genetically heterogeneous species. Its broad host range includes parrots and many other birds, but occasionally also humans (via zoonotic transmission), ruminants, horses, swine and rodents. To assess whether there are genetic markers associated with host tropism we comparatively analyzed whole-genome sequences of 61 C. psittaci strains, 47 of which carrying a 7.6-kbp plasmid. RESULTS: Following clean-up, reassembly and polishing of poorly assembled genomes from public databases, phylogenetic analyses using C. psittaci whole-genome sequence alignment revealed four major clades within this species. Clade 1 represents the most recent lineage comprising 40/61 strains and contains 9/10 of the psittacine strains, including type strain 6BC, and 10/13 of human isolates. Strains from different non-psittacine hosts clustered in Clades 2- 4. We found that clade membership correlates with typing schemes based on SNP types, ompA genotypes, multilocus sequence types as well as plasticity zone (PZ) structure and host preference. Genome analysis also revealed that i) sequence variation in the major outer membrane porin MOMP can result in 3D structural changes of immunogenic domains, ii) past host change of Clade 3 and 4 strains could be associated with loss of MAC/perforin in the PZ, rather than the large cytotoxin, iii) the distinct phylogeny of atypical strains (Clades 3 and 4) is also reflected in their repertoire of inclusion proteins (Inc family) and polymorphic membrane proteins (Pmps). CONCLUSIONS: Our study identified a number of genomic features that can be correlated with the phylogeny and host preference of C. psittaci strains. Our data show that intra-species genomic divergence is associated with past host change and includes deletions in the plasticity zone, structural variations in immunogenic domains and distinct repertoires of virulence factors.

BACKGROUND: Recent advances in CRISPR technology have enabled us to perform gene knock-in in various species and cell lines. CRISPR-mediated knock-in requires donor DNA which serves as a template for homology-directed repair (HDR). For knock-in of short sequences or base substitutions, ssDNA donors are frequently used among various other forms of HDR donors, such as linear dsDNA. However, partly due to the complexity of long ssDNA preparation, it remains unclear whether ssDNA is the optimal type of HDR donors for insertion of long transgenes such as fluorescent reporters in human cells. RESULTS: In this study, we established a nuclease-based simple method for the preparation of long ssDNA with high yield and purity, and comprehensively compared the performance of ssDNA and dsDNA donors with 90 bases of homology arms for endogenous gene tagging with long transgenes in human diploid RPE1 and HCT116 cells. Quantification using flow cytometry revealed lower efficiency of endogenous fluorescent tagging with ssDNA donors than with dsDNA. By analyzing knock-in outcomes using long-read amplicon sequencing and a classification framework, a variety of mis-integration events were detected regardless of the donor type. Importantly, the ratio of precise insertion was lower with ssDNA donors than with dsDNA. Moreover, in off-target integration analyses using donors without homology arms, ssDNA and dsDNA were comparably prone to non-homologous integration. CONCLUSIONS: These results indicate that ssDNA is not superior to dsDNA as long HDR donors with relatively short homology arms for gene knock-in in human RPE1 and HCT116 cells.

BACKGROUND: DNA methylation is one of the most abundant epigenetic modifications, which plays important roles in flower development, sex differentiation, and regulation of flowering time. Its pattern is affected by cytosine-5 DNA methyltransferase (C5-MTase) and DNA demethylase (dMTase). At present, there are no reports on C5-MTase and dMTase genes in heterodichogamous Cyclocarya paliurus. RESULTS: In this study, 6 CpC5-MTase and 3 CpdMTase genes were identified in diploid (2n = 2 ×  = 32) C. paliurus, while 20 CpC5-MTase and 13 CpdMTase genes were identified in autotetraploid (2n = 4 ×  = 64). 80% of identified genes maintained relatively fixed positions on chromosomes during polyploidization. In addition, we found that some DRM subfamily members didn't contain the UBA domain. The transcript abundance of CpC5-MTase and CpdMTase in male and female flowers of two morphs (protandry and protogyny) from diploidy was analyzed. Results showed that all genes were significantly up-regulated at the stage of floral bud break (S2), but significantly down-regulated at the stage of flower maturation (S4). At S2, some CpC5-MTase genes showed higher expression levels in PG-M than in PG-F, whereas some CpdMTase genes showed higher expression levels in PA-M than in PA-F. In addition, these genes were significantly associated with gibberellin synthesis-related genes (e.g. DELLA and GID1), suggesting that DNA methylation may play a role in the asynchronous floral development process through gibberellin signal. CONCLUSIONS: These results broaden our understanding of the CpC5-MTase and CpdMTase genes in diploid and autotetraploid C. paliurus, and provide a novel insight into regulatory mechanisms of DNA methylation in heterodichogamy.

Overexploitation of natural resources and pollution of seas, acidification of the ocean, and rising temperatures all contribute to the destruction of marine habitats and, in 2015, the protection of the ocean became one of the UN Sustainable Development Goals (SDG 14: Life Below Water). This collection aims to highlight the molecular genetic changes currently happening in marine organisms.

BACKGROUND: Testis is the only organ supporting sperm production and with the largest number of proteins and tissue-specific proteins in animals. In our previous studies, we have found that knockdown of ocnus (ocn), a testis-specific gene, resulted in much smaller testis with no germ cells in Drosophila melanogaster. However, the molecular consequences of ocn knockdown in fly testes are unknown. RESULTS: In this study, through iTRAQ quantitative proteomics sequencing, 606 proteins were identified from fly abdomens as having a significant and at least a 1.5-fold change in expression after ocn knockdown in fly testes, of which 85 were up-regulated and 521 were down-regulated. Among the differential expressed proteins (DEPs), apart from those proteins involved in spermatogenesis, the others extensively affected biological processes of generation of precursor metabolites and energy, metabolic process, and mitochondrial transport. Protein-protein interaction (PPI) analyses of DEPs showed that several kinases and/or phosphatases interacted with Ocn. Re-analyses of the transcriptome revealed 150 differential expressed genes (DEGs) appeared in the DEPs, and their changing trends in expressions after ocn knockdown were consistent. Many common down-regulated DEGs and DEPs were testis-specific or highly expressed in the testis of D. melanogaster. Quantitative RT-PCR (qRT-PCR) confirmed 12 genes appeared in both DEGs and DEPs were significantly down-regulated after ocn knockdown in fly testes. Furthermore, 153 differentially expressed phosphoproteins (DEPPs), including 72 up-regulated and 94 down-regulated phosphorylated proteins were also identified (13 phosphoproteins appeared in both up- and down-regulated groups due to having multiple phosphorylation sites). In addition to those DEPPs associated with spermatogenesis, the other DEPPs were enriched in actin filament-based process, protein folding, and mesoderm development. Some DEPs and DEPPs were involved in Notch, JAK/STAT, and cell death pathways. CONCLUSIONS: Given the drastic effect of the ocn knockdown on tissue development and testis cells composition, the differences in protein abundance in the ocn knockdown flies might not necessarily be the direct result of differential gene regulation due to the inactivation of ocn. Nevertheless, our results suggest that the expression of ocn is essential for Drosophila testis development and that its down-regulation disturbs key signaling pathways related to cell survival and differentiation. These DEPs and DEPPs identified may provide significant candidate set for future studies on the mechanism of male reproduction of animals, including humans.

BACKGROUND: The genetic locus responsible for duck body size has been fully explained before, but the growth trait-related genetic basis is still waiting to be explored. For example, the genetic site related to growth rate, an important economic trait affecting marketing weight and feeding cost, is still unclear. Here, we performed genome wide association study (GWAS) to identify growth rate-associated genes and mutations. RESULT: In the current study, the body weight data of 358 ducks were recorded every 10 days from hatching to 120 days of age. According to the growth curve, we evaluated the relative and absolute growth rates (RGR and AGR) of 5 stages during the early rapid growth period. GWAS results for RGRs identified 31 significant SNPs on autosomes, and these SNPs were annotated by 24 protein-coding genes. Fourteen autosomal SNPs were significantly associated with AGRs. In addition, 4 shared significant SNPs were identified as having an association with both AGR and RGR, which were Chr2: 11483045 C>T, Chr2: 13750217 G>A, Chr2: 42508231 G>A and Chr2: 43644612 C>T. Among them, Chr2: 11483045 C>T, Chr2: 42508231 G>A, and Chr2: 43644612 C>T were annotated by ASAP1, LYN and CABYR, respectively. ASAP1 and LYN have already been proven to play roles in the growth and development of other species. In addition, we genotyped every duck using the most significant SNP (Chr2: 42508231 G>A) and compared the growth rate difference among each genotype population. The results showed that the growth rates of individuals carrying the Chr2: 42508231 A allele were significantly lower than those without this allele. Moreover, the results of the Mendelian randomization (MR) analysis supported the idea that the growth rate and birth weight had a causal effect on the adult body weight, with the growth rate having a greater effect size. CONCLUSION: In this study, 41 SNPs significantly related to growth rate were identified. In addition, we considered that the ASAP1 and LYN genes are essential candidate genes affecting the duck growth rate. The growth rate also showed the potential to be used as a reliable predictor of adult weight, providing a theoretical reference for preselection.

BACKGROUND: The genus Acidithiobacillus has been widely concerned due to its superior survival and oxidation ability in acid mine drainage (AMD). However, the contribution of insertion sequence (IS) to their biological evolution and environmental adaptation is very limited. ISs are the simplest kinds of mobile genetic elements (MGEs), capable of interrupting genes, operons, or regulating the expression of genes through transposition activity. ISs could be classified into different families with their own members, possessing different copies. RESULTS: In this study, the distribution and evolution of ISs, as well as the functions of the genes around ISs in 36 Acidithiobacillus genomes, were analyzed. The results showed that 248 members belonging to 23 IS families with a total of 10,652 copies were identified within the target genomes. The IS families and copy numbers among each species were significantly different, indicating that the IS distribution of Acidithiobacillus were not even. A. ferrooxidans had 166 IS members, which may develop more gene transposition strategies compared with other Acidithiobacillus spp. What's more, A. thiooxidans harbored the most IS copies, suggesting that their ISs were the most active and more likely to transpose. The ISs clustered in the phylogenetic tree approximately according to the family, which were mostly different from the evolutionary trends of their host genomes. Thus, it was suggested that the recent activity of ISs of Acidithiobacillus was not only determined by their genetic characteristics, but related with the environmental pressure. In addition, many ISs especially Tn3 and IS110 families were inserted around the regions whose functions were As/Hg/Cu/Co/Zn/Cd translocation and sulfur oxidation, implying that ISs could improve the adaptive capacities of Acidithiobacillus to the extremely acidic environment by enhancing their resistance to heavy metals and utilization of sulfur. CONCLUSIONS: This study provided the genomic evidence for the contribution of IS to evolution and adaptation of Acidithiobacillus, opening novel sights into the genome plasticity of those acidophiles.

BACKGROUND: Piwi-interacting RNAs (piRNAs) have been proven to be closely associated with human diseases. The identification of the potential associations between piRNA and disease is of great significance for complex diseases. Traditional "wet experiment" is time-consuming and high-priced, predicting the piRNA-disease associations by computational methods is of great significance. METHODS: In this paper, a method based on the embedding transformation graph convolution network is proposed to predict the piRNA-disease associations, named ETGPDA. Specifically, a heterogeneous network is constructed based on the similarity information of piRNA and disease, as well as the known piRNA-disease associations, which is applied to extract low-dimensional embeddings of piRNA and disease based on graph convolutional network with an attention mechanism. Furthermore, the embedding transformation module is developed for the problem of embedding space inconsistency, which is lightweighter, stronger learning ability and higher accuracy. Finally, the piRNA-disease association score is calculated by the similarity of the piRNA and disease embedding. RESULTS: Evaluated by fivefold cross-validation, the AUC of ETGPDA achieves 0.9603, which is better than the other five selected computational models. The case studies based on Head and neck squamous cell carcinoma and Alzheimer's disease further prove the superior performance of ETGPDA. CONCLUSIONS: Hence, the ETGPDA is an effective method for predicting the hidden piRNA-disease associations.

BACKGROUND: Cell clustering is a prerequisite for identifying differentially expressed genes (DEGs) in single-cell RNA sequencing (scRNA-seq) data. Obtaining a perfect clustering result is of central importance for subsequent analyses, but not easy. Additionally, the increase in cell throughput due to the advancement of scRNA-seq protocols exacerbates many computational issues, especially regarding method runtime. To address these difficulties, a new, accurate, and fast method for detecting DEGs in scRNA-seq data is needed. RESULTS: Here, we propose single-cell minimum enclosing ball (scMEB), a novel and fast method for detecting single-cell DEGs without prior cell clustering results. The proposed method utilizes a small part of known non-DEGs (stably expressed genes) to build a minimum enclosing ball and defines the DEGs based on the distance of a mapped gene to the center of the hypersphere in a feature space. CONCLUSIONS: We compared scMEB to two different approaches that could be used to identify DEGs without cell clustering. The investigation of 11 real datasets revealed that scMEB outperformed rival methods in terms of cell clustering, predicting genes with biological functions, and identifying marker genes. Moreover, scMEB was much faster than the other methods, making it particularly effective for finding DEGs in high-throughput scRNA-seq data. We have developed a package scMEB for the proposed method, which could be available at https://github.com/FocusPaka/scMEB .

BACKGROUND: Cyclic nucleotide-gated ion channels (CNGCs) are nonselective cation channels that are ubiquitous in eukaryotic organisms. As Ca2+ channels, some CNGCs have also proven to be K+-permeable and involved in plant development and responses to environmental stimuli. Sugarcane is an important sugar and energy crop worldwide. However, reports on CNGC genes in sugarcane are limited. RESULTS: In this study, 16 CNGC genes and their alleles were identified from Saccharum spontaneum and classified into 5 groups based on phylogenetic analysis. Investigation of gene duplication and syntenic relationships between S. spontaneum and both rice and Arabidopsis demonstrated that the CNGC gene family in S. spontaneum expanded primarily by segmental duplication events. Many SsCNGCs showed variable expression during growth and development as well as in tissues, suggesting functional divergence. Light-responsive cis-acting elements were discovered in the promoters of all the identified SsCNGCs, and the expression of most of the SsCNGCs showed a diurnal rhythm. In sugarcane, the expression of some SsCNGCs was regulated by low-K+ treatment. Notably, SsCNGC13 may be involved in both sugarcane development and its response to environmental stimuli, including response to low-K+ stress. CONCLUSION: This study identified the CNGC genes in S. spontaneum and provided insights into the transcriptional regulation of these SsCNGCs during development, circadian rhythm and under low-K+ stress. These findings lay a theoretical foundation for future investigations of the CNGC gene family in sugarcane.

BACKGROUND: Macleaya cordata is a traditional medicinal herb, and it has high tolerance and accumulation ability to heavy metals, which make it a good candidate species for studying phytoremediation. The objectives of this study were to investigate response and tolerance of M. cordata to lead (Pb) toxicity based on comparative analysis of transcriptome and proteome. RESULTS: In this study, the seedlings of M. cordata cultured in Hoagland solution were treated with 100 µmol·L- 1 Pb for 1 day (Pb 1d) or 7 days (Pb 7d), subsequently leaves of M. cordata were taken for the determination of Pb accumulation and hydrogen peroxide production (H2O2), meanwhile a total number of 223 significantly differentially expressed genes (DEGs) and 296 differentially expressed proteins (DEPs) were screened between control and Pb treatments. The results showed leaves of M. cordata had a special mechanism to maintain Pb at an appropriate level. Firstly, some DEGs were iron (Fe) deficiency-induced transporters, for example, genes of vacuolar iron transporter and three ABC transporter I family numbers were upregulated by Pb, which can maintain Fe homeostasis in cytoplasm or chloroplast. In addition, five genes of calcium (Ca2+) binding proteins were downregulated in Pb 1d, which may regulate cytoplasmic Ca2+ concentration and H2O2 signaling pathway. On the other hand, the cysteine synthase upregulated, glutathione S-transferase downregulated and glutathione reductase downregulated in Pb 7d can cause reduced glutathione accumulation and decrease Pb detoxification in leaves. Furthermore, DEPs of eight chlorophyll a/b binding proteins, five ATPases and eight ribosomal proteins can play a pivotal role on chloroplast turnover and ATP metabolism. CONCLUSIONS: Our results suggest that the proteins involved in Fe homeostasis and chloroplast turnover in mesophyll cells may play key roles in tolerance of M. cordata to Pb. This study offers some novel insights into Pb tolerance mechanism of plants, and the potential valuable for environmental remediation of this important medicinal plant.

BACKGROUND: Despite being in the 21st century, the world has still not been able to vanquish the global AIDS epidemic, and the only foreseeable solution seems to be a safe and effective vaccine. Unfortunately, vaccine trials so far have returned unfruitful results, possibly due to their inability to induce effective cellular, humoral and innate immune responses. The current study aims to tackle these limitations and propose the desired vaccine utilizing immunoinformatic approaches that have returned promising results in designing vaccines against various rapidly mutating organisms. For this, all polyprotein and protein sequences of HIV-1 were retrieved from the LANL (Los Alamos National Laboratory) database. The consensus sequence was generated after alignment and used to predict epitopes. Conserved, antigenic, non-allergenic, T-cell inducing, B-cell inducing, IFN-ɣ inducing, non-human homologous epitopes were selected and combined to propose two vaccine constructs i.e., HIV-1a (without adjuvant) and HIV-1b (with adjuvant). RESULTS: HIV-1a and HIV-1b were subjected to antigenicity, allergenicity, structural quality analysis, immune simulations, and MD (molecular dynamics) simulations. Both proposed multi-epitope vaccines were found to be antigenic, non-allergenic, stable, and induce cellular, humoral, and innate immune responses. TLR-3 docking and in-silico cloning of both constructs were also performed. CONCLUSION: Our results indicate HIV-1b to be more promising than HIV-1a; experimental validations can confirm the efficacy and safety of both constructs and in-vivo efficacy in animal models.

Apicomplexa are ancient and diverse organisms which have been poorly characterized by modern genomics. To better understand the evolution and diversity of these single-celled eukaryotes, we sequenced the genome of Ophryocystis elektroscirrha, a parasite of monarch butterflies, Danaus plexippus. We contextualize our newly generated resources within apicomplexan genomics before answering longstanding questions specific to this host-parasite system. To start, the genome is miniscule, totaling only 9 million bases and containing fewer than 3,000 genes, half the gene content of two other sequenced invertebrate-infecting apicomplexans, Porospora gigantea and Gregarina niphandrodes. We found that O. elektroscirrha shares different orthologs with each sequenced relative, suggesting the true set of universally conserved apicomplexan genes is very small indeed. Next, we show that sequencing data from other potential host butterflies can be used to diagnose infection status as well as to study diversity of parasite sequences. We recovered a similarly sized parasite genome from another butterfly, Danaus chrysippus, that was highly diverged from the O. elektroscirrha reference, possibly representing a distinct species. Using these two new genomes, we investigated potential evolutionary response by parasites to toxic phytochemicals their hosts ingest and sequester. Monarch butterflies are well-known to tolerate toxic cardenolides thanks to changes in the sequence of their Type II ATPase sodium pumps. We show that Ophryocystis completely lacks Type II or Type 4 sodium pumps, and related proteins PMCA calcium pumps show extreme sequence divergence compared to other Apicomplexa, demonstrating new avenues of research opened by genome sequencing of non-model Apicomplexa.

BACKGROUND: Armillaria species are plant pathogens, but a few Armillaria species can establish a symbiotic relationship with Gastrodia elata, a rootless and leafless orchid, that is used as a Chinese herbal medicine. Armillaria is a nutrient source for the growth of G. elata. However, there are few reports on the molecular mechanism of symbiosis between Armillaria species and G. elata. The genome sequencing and analysis of Armillaria symbiotic with G. elata would provide genomic information for further studying the molecular mechanism of symbiosis. RESULTS: The de novo genome assembly was performed with the PacBio Sequel platform and Illumina NovaSeq PE150 for the A. gallica Jzi34 strain, which was symbiotic with G. elata. Its genome assembly contained ~ 79.9 Mbp and consisted of 60 contigs with an N50 of 2,535,910 bp. There were only 4.1% repetitive sequences in the genome assembly. Functional annotation analysis revealed a total of 16,280 protein coding genes. Compared with the other five genomes of Armillaria, the carbohydrate enzyme gene family of the genome was significantly contracted, while it had the largest set of glycosyl transferase (GT) genes. It also had an expansion of auxiliary activity enzymes AA3-2 gene subfamily and cytochrome P450 genes. The synteny analysis result of P450 genes reveals that the evolutionary relationship of P450 proteins between A. gallica Jzi34 and other four Armillaria was complex. CONCLUSIONS: These characteristics may be beneficial for establishing a symbiotic relationship with G. elata. These results explore the characteristics of A. gallica Jzi34 from a genomic perspective and provide an important genomic resource for further detailed study of Armillaria. This will help to further study the symbiotic mechanism between A. gallica and G. elata.

The risk of exposure of the general public or military personnel to high levels of ionizing radiation from nuclear weapons or radiological accidents is a dire national security matter. The development of advanced molecular biodosimetry methods, those that measure biological response, such as transcriptomics, to screen large populations of radiation-exposed victims is key to improving survival outcomes during radiological mass casualty scenarios. In this study, nonhuman primates were exposed to either 12.0 Gy cobalt-60 gamma (total-body irradiation, TBI) or X-ray (partial-body irradiation, PBI) 24 h after administration of a potential radiation medical countermeasure, gamma-tocotrienol (GT3). Changes in the jejunal transcriptomic profiles in GT3-treated and irradiated animals were compared to healthy controls to assess the extent of radiation damage. No major effect of GT3 on radiation-induced transcriptome at this radiation dose was identified. About 80% of the pathways with a known activation or repression state were commonly observed between both exposures. Several common pathways activated due to irradiation include FAK signaling, CREB signaling in the neurons, phagosome formation, and G-protein coupled signaling pathway. Sex-specific differences associated with excessive mortality among irradiated females were identified in this study, including Estrogen receptor signaling. Differential pathway activation was also identified across PBI and TBI, pointing towards altered molecular response for different degrees of bone marrow sparing and radiation doses. This study provides insight into radiation-induced changes in jejunal transcriptional profiles, supporting the investigation for the identification of biomarkers for radiation injury and countermeasure efficacy.

BACKGROUND: Single-cell RNA sequencing is a state-of-the-art technology to understand gene expression in complex tissues. With the growing amount of data being generated, the standardization and automation of data analysis are critical to generating hypotheses and discovering biological insights. RESULTS: Here, we present scRNASequest, a semi-automated single-cell RNA-seq (scRNA-seq) data analysis workflow which allows (1) preprocessing from raw UMI count data, (2) harmonization by one or multiple methods, (3) reference-dataset-based cell type label transfer and embedding projection, (4) multi-sample, multi-condition single-cell level differential gene expression analysis, and (5) seamless integration with cellxgene VIP for visualization and with CellDepot for data hosting and sharing by generating compatible h5ad files. CONCLUSIONS: We developed scRNASequest, an end-to-end pipeline for single-cell RNA-seq data analysis, visualization, and publishing. The source code under MIT open-source license is provided at https://github.com/interactivereport/scRNASequest . We also prepared a bookdown tutorial for the installation and detailed usage of the pipeline: https://interactivereport.github.io/scRNAsequest/tutorial/docs/ . Users have the option to run it on a local computer with a Linux/Unix system including MacOS, or interact with SGE/Slurm schedulers on high-performance computing (HPC) clusters.

BACKGROUND: It is now possible to analyze cellular heterogeneity at the single-cell level thanks to the rapid developments in single-cell sequencing technologies. The clustering of cells is a fundamental and common step in heterogeneity analysis. Even so, accurate cell clustering remains a challenge due to the high levels of noise, the high dimensions, and the high sparsity of data. RESULTS: Here, we present SCEA, a clustering approach for scRNA-seq data. Using two consecutive units, an encoder based on MLP and a graph attention auto-encoder, to obtain cell embedding and gene embedding, SCEA can simultaneously achieve cell low-dimensional representation and clustering performing various examinations to obtain the optimal value for each parameter, the presented result is in its most optimal form. To evaluate the performance of SCEA, we performed it on several real scRNA-seq datasets for clustering and visualization analysis. CONCLUSIONS: The experimental results show that SCEA generally outperforms several popular single-cell analysis methods. As a result of using all available datasets, SCEA, in average, improves clustering accuracy by 4.4% in ARI Parameters over the well-known method scGAC. Also, the accuracy improvement of 11.65% is achieved by SCEA, compared to the Seurat model.

BACKGROUND: Mitochondrial genome sequences have become critical to the study of biodiversity. Genome skimming and other short-read based methods are the most common approaches, but they are not well-suited to scale up to multiplexing hundreds of samples. Here, we report on a new approach to sequence hundreds to thousands of complete mitochondrial genomes in parallel using long-amplicon sequencing. We amplified the mitochondrial genome of 677 specimens in two partially overlapping amplicons and implemented an asymmetric PCR-based indexing approach to multiplex 1,159 long amplicons together on a single PacBio SMRT Sequel II cell. We also tested this method on Oxford Nanopore Technologies (ONT) MinION R9.4 to assess if this method could be applied to other long-read technologies. We implemented several optimizations that make this method significantly more efficient than alternative mitochondrial genome sequencing methods. RESULTS: With the PacBio sequencing data we recovered at least one of the two fragments for 96% of samples (~ 80-90%) with mean coverage ~ 1,500x. The ONT data recovered less than 50% of input fragments likely due to low throughput and the design of the Barcoded Universal Primers which were optimized for PacBio sequencing. We compared a single mitochondrial gene alignment to half and full mitochondrial genomes and found, as expected, increased tree support with longer alignments, though whole mitochondrial genomes were not significantly better than half mitochondrial genomes. CONCLUSIONS: This method can effectively capture thousands of long amplicons in a single run and be used to build more robust phylogenies quickly and effectively. We provide several recommendations for future users depending on the evolutionary scale of their system. A natural extension of this method is to collect multi-locus datasets consisting of mitochondrial genomes and several long nuclear loci at once.

BACKGROUND: The molecular mechanisms underlying the onset and progression of irreversible pulpitis have been studied for decades. Many studies have indicated a potential correlation between autophagy and this disease. Against the background of the competing endogenous RNA (ceRNA) theory, protein-coding RNA functions are linked with long noncoding RNAs (lncRNAs) and microRNAs (miRNAs). This mechanism has been widely studied in various fields but has rarely been reported in the context of irreversible pulpitis. The hub genes selected under this theory may represent the key to the interaction between autophagy and irreversible pulpitis. RESULTS: Filtering and differential expression analyses of the GSE92681 dataset, which contains data from 7 inflamed and 5 healthy pulp tissue samples, were conducted. The results were intersected with autophagy-related genes (ARGs), and 36 differentially expressed ARGs (DE-ARGs) were identified. Functional enrichment analysis and construction of the protein‒protein interaction (PPI) network of DE-ARGs were performed. Coexpression analysis was conducted between differentially expressed lncRNAs (DElncRNAs) and DE-ARGs, and 151 downregulated and 59 upregulated autophagy-related DElncRNAs (AR-DElncRNAs) were identified. StarBase and multiMiR were then used to predict related microRNAs of AR-DElncRNAs and DE-ARGs, respectively. We established ceRNA networks including 9 hub lncRNAs (HCP5 and AC112496.1 ↑; FENDRR, AC099850.1, ZSWIM8-AS1, DLX6-AS1, LAMTOR5-AS1, TMEM161B-AS1 and AC145207.5 ↓), which were validated by a qRT‒PCR analysis of pulp tissue from patients with irreversible pulpitis. CONCLUSION: We constructed two networks consisting of 9 hub lncRNAs based on the comprehensive identification of autophagy-related ceRNAs. This study may provide novel insights into the interactive relationship between autophagy and irreversible pulpitis and identifies several lncRNAs that may serve as potential biological markers.

Macrophages are important effector cells in tumor progression and immune regulation. Previously, we demonstrated that the transcription suppressor homeobox containing 1(HMBOX1) exhibits immunosuppressive activity in LPS-induced acute liver injury by impeding macrophage infiltration and activation. We also observed a lower proliferation in HMBOX1-overexpressed RAW264.7 cells. However, the specific mechanism was unclear. Here, a work was performed to characterize HMBOX1 function related to cell proliferation from a metabolomics standpoint by comparing the metabolic profiles of HMBOX1-overexpressed RAW264.7 cells to those of the controls. Firstly, we assessed HMBOX1 anti-proliferation activity in RAW264.7 cells with CCK8 assay and clone formation. Then, we performed metabolomic analyses by ultra-liquid chromatography coupled with mass spectrometry to explore the potential mechanisms. Our results indicated that HMBOX1 inhibited the macrophage growth curve and clone formation ability. Metabolomic analyses showed significant changes in HMBOX1-overexpressed RAW264.7 metabolites. A total of 1312 metabolites were detected, and 185 differential metabolites were identified based on the criterion of OPLS-DA VIP > 1 and p value < 0.05. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that the elevated HMBOX1 in RAW264.7 inhibited the pathways of amino acid and nucleotide metabolism. Glutamine concentrations decreased significantly in HMBOX1-overexpressed macrophages, and glutamine-related transporter SLC1A5 was also downregulated. Furthermore, SLC1A5 overexpression reversed HMBOX1 inhibition of macrophage proliferation. This study demonstrated the potential mechanism of the HMBOX1/SLC1A5 pathway in cell proliferation by regulating glutamine transportation. The results may help provide a new direction for therapeutic interventions in macrophage-related inflammatory diseases.

BACKGROUND: Ilex pubescens is an important traditional Chinese medicinal plant with many naturally occurring compounds and multiple pharmacological effects. However, the lack of reference genomic information has led to tardiness in molecular biology research and breeding programs of this plant. RESULTS: To obtain knowledge on the genomic information of I. pubescens, a genome survey was performed for the first time by next generation sequencing (NGS) together with genome size estimation using flow cytometry. The whole genome survey of I. pubescens generated 46.472 Gb of sequence data with approximately 82.2 × coverage. K-mer analysis indicated that I. pubescens has a small genome of approximately 553 Mb with 1.93% heterozygosity rate and 39.1% repeat rate. Meanwhile, the genome size was estimated to be 722 Mb using flow cytometry, which was possibly more precise for assessment of genome size than k-mer analysis. A total of 45.842 Gb clean reads were assembled into 808,938 scaffolds with a relatively short N50 of 760 bp. The average guanine and cytosine (GC) content was 37.52%. In total, 197,429 microsatellite motifs were detected with a frequency of 2.8 kb, among which mononucleotide motifs were the most abundant (up to 62.47% of the total microsatellite motifs), followed by dinucleotide and trinucleotide motifs. CONCLUSION: In summary, the genome of I. pubescens is small but complex with a high level of heterozygosity. Even though not successfully applied for estimation of genome size due to its complex genome, the survey sequences will help to design whole genome sequencing strategies and provide genetic information support for resource protection, genetic diversity analysis, genetic improvement and artificial breeding of I. pubescens.

BACKGROUND: Seagull as a migratory wild bird has become most popular species in southwest China since 1980s. Previously, we analyzed the gut microbiota and intestinal pathogenic bacteria configuration for this species by using 16S rRNA sequencing and culture methods. To continue in-depth research on the gut microbiome of migratory seagulls, the metagenomics, DNA virome and RNA virome were both investigated for their gut microbial communities of abundance and diversity in this study. RESULTS: The metagenomics results showed 99.72% of total species was bacteria, followed by viruses, fungi, archaea and eukaryota. In particular, Shigella sonnei, Escherichia albertii, Klebsiella pneumonia, Salmonella enterica and Shigella flexneri were the top distributed taxa at species level. PCoA, NMDS, and statistics indicated some drug resistant genes, such as adeL, evgS, tetA, PmrF, and evgA accumulated as time went by from November to January of the next year, and most of these genes were antibiotic efflux. DNA virome composition demonstrated that Caudovirales was the most abundance virus, followed by Cirlivirales, Geplafuvirales, Petitvirales and Piccovirales. Most of these phages corresponded to Enterobacteriaceae and Campylobacteriaceae bacterial hosts respectively. Caliciviridae, Coronaviridae and Picornaviridae were the top distributed RNA virome at family level of this migratory animal. Phylogenetic analysis indicated the sequences of contigs of Gammacoronavirus and Deltacoronavirus had highly similarity with some coronavirus references. CONCLUSIONS: In general, the characteristics of gut microbiome of migratory seagulls were closely related to human activities, and multiomics still revealed the potential public risk to human health.

BACKGROUND: To reduce the cost of genomic selection, a low-density (LD) single nucleotide polymorphism (SNP) chip can be used in combination with imputation for genotyping selection candidates instead of using a high-density (HD) SNP chip. Next-generation sequencing (NGS) techniques have been increasingly used in livestock species but remain expensive for routine use for genomic selection. An alternative and cost-efficient solution is to use restriction site-associated DNA sequencing (RADseq) techniques to sequence only a fraction of the genome using restriction enzymes. From this perspective, use of RADseq techniques followed by an imputation step on HD chip as alternatives to LD chips for genomic selection was studied in a pure layer line. RESULTS: Genome reduction and sequencing fragments were identified on reference genome using four restriction enzymes (EcoRI, TaqI, AvaII and PstI) and a double-digest RADseq (ddRADseq) method (TaqI-PstI). The SNPs contained in these fragments were detected from the 20X sequence data of the individuals in our population. Imputation accuracy on HD chip with these genotypes was assessed as the mean correlation between true and imputed genotypes. Several production traits were evaluated using single-step GBLUP methodology. The impact of imputation errors on the ranking of the selection candidates was assessed by comparing a genomic evaluation based on ancestry using true HD or imputed HD genotyping. The relative accuracy of genomic estimated breeding values (GEBVs) was investigated by considering the GEBVs estimated on offspring as a reference. With AvaII or PstI and ddRADseq with TaqI and PstI, more than 10 K SNPs were detected in common with the HD SNP chip, resulting in an imputation accuracy greater than 0.97. The impact of imputation errors on genomic evaluation of the breeders was reduced, with a Spearman correlation greater than 0.99. Finally, the relative accuracy of GEBVs was equivalent. CONCLUSIONS: RADseq approaches can be interesting alternatives to low-density SNP chips for genomic selection. With more than 10 K SNPs in common with the SNPs of the HD SNP chip, good imputation and genomic evaluation results can be obtained. However, with real data, heterogeneity between individuals with missing data must be considered.

BACKGROUND: Oriental river prawn Macrobrachium nipponense is an economically important aquaculture species in China, Japan, and Vietnam. In commercial prawn farming, feed cost constitutes about 50 to 65% of the actual variable cost. Improving feed conversion efficiency in prawn culture will not only increase economic benefit, but also save food and protect the environment. The common indicators used for feed conversion efficiency include feed conversion ratio (FCR), feed efficiency ratio (FER), and residual feed intake (RFI). Among these, RFI is much more suitable than FCR and FER during the genetic improvement of feed conversion efficiency for aquaculture species. RESULTS: In this study, the transcriptome and metabolome of hepatopancreas and muscle of M. nipponense from high RFI low RFI groups, which identified after culture for 75 days, were characterized using combined transcriptomic and metabolomic analysis. A total of 4540 differentially expressed genes (DEGs) in hepatopancreas, and 3894 DEGs in muscle were identified, respectively. The DEGs in hepatopancreas were mainly enriched in KEGG pathways including the metabolism of xenobiotics by cytochrome P450 (down-regulated), fat digestion and absorption (down-regulated) and aminoacyl-tRNA biosynthesis (up-regulated), etc. The DEGs in muscle were mainly enriched in KEGG pathways including the protein digestion and absorption (down-regulated), glycolysis/gluconeogenesis (down-regulated), and glutathione metabolism (up-regulated), etc. At the transcriptome level, the RFI of M. nipponense was mainly controlled in biological pathways such as the high immune expression and the reduction of nutrients absorption capacity. A total of 445 and 247 differently expressed metabolites (DEMs) were identified in the hepatopancreas and muscle, respectively. At the metabolome level, the RFI of M. nipponense was affected considerably by amino acid and lipid metabolism. CONCLUSIONS: M. nipponense from higher and lower RFI groups have various physiological and metabolic capability processes. The down-regulated genes, such as carboxypeptidase A1, 6-phosphofructokinase, long-chain-acyl-CoA dehydrogenase, et. al., in digestion and absorption of nutrients, and the up-regulated metabolites, such as aspirin, lysine, et. al., in response to immunity could be potential candidate factors contributed to RFI variation for M. nipponense. Overall, these results would provide new insights into the molecular mechanism of feed conversion efficiency and assist in selective breeding to improve feed conversion efficiency in M. nipponense.

BACKGROUND: Cattle (Bos taurus) are a major large livestock, however, compared with other species, the transcriptional specificity of bovine oocyte development has not been emphasised. RESULTS: To reveal the unique transcriptional signatures of bovine oocyte development, we used integrated multispecies comparative analysis and weighted gene co-expression network analysis (WGCNA) to perform bioinformatic analysis of the germinal follicle (GV) and second meiosis (MII) gene expression profile from cattle, sheep, pigs and mice. We found that the expression levels of most genes were down-regulated from GV to MII in all species. Next, the multispecies comparative analysis showed more genes involved in the regulation of cAMP signalling during bovine oocyte development. Moreover, the green module identified by WGCNA was closely related to bovine oocyte development. Finally, integrated multispecies comparative analysis and WGCNA picked up 61 bovine-specific signature genes that participate in metabolic regulation and steroid hormone biosynthesis. CONCLUSION: In a short, this study provides new insights into the regulation of cattle oocyte development from a cross-species comparison.

BACKGROUND: The prevalence of the COVID-19 disease in recent years and its widespread impact on mortality, as well as various aspects of life around the world, has made it important to study this disease and its viral cause. However, very long sequences of this virus increase the processing time, complexity of calculation, and memory consumption required by the available tools to compare and analyze the sequences. RESULTS: We present a new encoding method, named PC-mer, based on the k-mer and physic-chemical properties of nucleotides. This method minimizes the size of encoded data by around 2 k times compared to the classical k-mer based profiling method. Moreover, using PC-mer, we designed two tools: 1) a machine-learning-based classification tool for coronavirus family members with the ability to recive input sequences from the NCBI database, and 2) an alignment-free computational comparison tool for calculating dissimilarity scores between coronaviruses at the genus and species levels. CONCLUSIONS: PC-mer achieves 100% accuracy despite the use of very simple classification algorithms based on Machine Learning. Assuming dynamic programming-based pairwise alignment as the ground truth approach, we achieved a degree of convergence of more than 98% for coronavirus genus-level sequences and 93% for SARS-CoV-2 sequences using PC-mer in the alignment-free classification method. This outperformance of PC-mer suggests that it can serve as a replacement for alignment-based approaches in certain sequence analysis applications that rely on similarity/dissimilarity scores, such as searching sequences, comparing sequences, and certain types of phylogenetic analysis methods that are based on sequence comparison.

Long non-coding RNAs (lncRNAs) play a crucial role in numbers of biological processes and have received wide attention during the past years. Since the rapid development of high-throughput transcriptome sequencing technologies (RNA-seq) lead to a large amount of RNA data, it is urgent to develop a fast and accurate coding potential predictor. Many computational methods have been proposed to address this issue, they usually exploit information on open reading frame (ORF), protein sequence, k-mer, evolutionary signatures, or homology. Despite the effectiveness of these approaches, there is still much room to improve. Indeed, none of these methods exploit the contextual information of RNA sequence, for example, k-mer features that counts the occurrence frequencies of continuous nucleotides (k-mer) in the whole RNA sequence cannot reflect local contextual information of each k-mer. In view of this shortcoming, here, we present a novel alignment-free method, CPPVec, which exploits the contextual information of RNA sequence for coding potential prediction for the first time, it can be easily implemented by distributed representation (e.g., doc2vec) of protein sequence translated from the longest ORF. The experimental findings demonstrate that CPPVec is an accurate coding potential predictor and significantly outperforms existing state-of-the-art methods.

BACKGROUND: The genus Daphnia switches its reproductive mode from subitaneous egg production to resting egg production in response to environmental stimuli. Although this life history trait is essential for surviving unsuitable environments, the molecular mechanism of resting egg production is little understood. In this study, we examined genes related to induction of resting egg production using two genotypes of panarctic Daphnia pulex, the JPN1 and JPN2 lineages, which differ genetically in the frequency of resting egg production. We reared these genotypes under high and low food levels. At the high food level, individuals of both genotypes continually produced subitaneous eggs, whereas at the low food level, only the JPN2 genotype produced resting eggs. Then, we performed RNA-seq analysis on specimens of three instars, including before and after egg production. RESULTS: These results showed that expressed genes differed significantly between individuals grown under high and low food levels and among individuals of different instars and genotypes. Among these differentially expressed genes (DEGs), we found 16 that changed expression level before resting egg production. Some of these genes showed high-level expression only before resting egg production and one gene was an ortholog of bubblegum (bgm), which is reportedly up-regulated before diapause in bumblebees. According to gene ontology (GO) enrichment analysis, one GO term annotated as long-chain fatty acid biosynthetic process was enriched among these 16 genes. In addition, GO terms related to glycometabolism were enriched among down-regulated genes of individuals holding resting eggs, compared to those before resting egg production. CONCLUSIONS: We found candidate genes highly expressed only before resting egg production. Although functions of candidate genes found in this study have not been reported previously in Daphnia, catabolism of long-chain fatty acids and metabolism of glycerates are related to diapause in other organisms. Thus, it is highly probable that candidate genes identified in this study are related to the molecular mechanism regulating resting egg production in Daphnia.

BACKGROUND: Huntiella resides in the Ceratocystidaceae, a family of fungi that accommodates important plant pathogens and insect-associated saprotrophs. Species in the genus have either heterothallic or unisexual (a form of homothallism) mating systems, providing an opportunity to investigate the genetic mechanisms that enable transitions between reproductive strategies in related species. Two newly sequenced Huntiella genomes are introduced in this study and comparative genomics and transcriptomics tools are used to investigate the differences between heterothallism and unisexuality across the genus. RESULTS: Heterothallic species harbored up to seven copies of the a-factor pheromone, each of which possessed numerous mature peptide repeats. In comparison, unisexual Huntiella species had only two or three copies of this gene, each with fewer repeats. Similarly, while the heterothallic species expressed up to 12 copies of the mature α-factor pheromone, unisexual species had up to six copies. These significant differences imply that unisexual Huntiella species do not rely on a mating partner recognition system in the same way that heterothallic fungi do. CONCLUSION: While it is suspected that mating type-independent pheromone expression is the mechanism allowing for unisexual reproduction in Huntiella species, our results suggest that the transition to unisexuality may also have been associated with changes in the genes governing the pheromone pathway. While these results are specifically related to Huntiella, they provide clues leading to a better understanding of sexual reproduction and the fluidity of mating strategies in fungi more broadly.

BACKGROUND: Human primary hepatocytes (PHCs) are considered to be the best cell source for cell-based therapies for the treatment of end-stage liver disease and acute liver failure. To obtain sufficient and high-quality functional human hepatocytes, we have established a strategy to dedifferentiate human PHCs into expandable hepatocyte-derived liver progenitor-like cells (HepLPCs) through in vitro chemical reprogramming. However, the reduced proliferative capacity of HepLPCs after long-term culture still limits their utility. Therefore, in this study, we attempted to explore the potential mechanism related to the proliferative ability of HepLPCs in vitro culture. RESULTS: In this study, analysis of assay for transposase accessible chromatin using sequencing (ATAC-seq) and RNA sequencing (RNA-seq) were performed for PHCs, proliferative HepLPCs (pro-HepLPCs) and late-passage HepLPCs (lp-HepLPCs). Genome-wide transcriptional and chromatin accessibility changes during the conversion and long-term culture of HepLPCs were studied. We found that lp-HepLPCs exhibited an aged phenotype characterized by the activation of inflammatory factors. Epigenetic changes were found to be consistent with our gene expression findings, with promoter and distal regions of many inflammatory-related genes showing increased accessibility in the lp-HepLPCs. FOSL2, a member of the AP-1 family, was found to be highly enriched in the distal regions with increased accessibility in lp-HepLPCs. Its depletion attenuated the expression of aging- and senescence-associated secretory phenotype (SASP)-related genes and resulted in a partial improvement of the aging phenotype in lp-HepLPCs. CONCLUSIONS: FOSL2 may drive the aging of HepLPCs by regulating inflammatory factors and its depletion may attenuate this phenotypic shift. This study provides a novel and promising approach for the long-term in vitro culture of HepLPCs.

BACKGROUND: Yellow or stripe rust, caused by the fungus Puccinia striiformis f. sp. tritici (Pst) is an important disease of wheat that threatens wheat production. Since developing resistant cultivars offers a viable solution for disease management, it is essential to understand the genetic basis of stripe rust resistance. In recent years, meta-QTL analysis of identified QTLs has gained popularity as a way to dissect the genetic architecture underpinning quantitative traits, including disease resistance. RESULTS: Systematic meta-QTL analysis involving 505 QTLs from 101 linkage-based interval mapping studies was conducted for stripe rust resistance in wheat. For this purpose, publicly available high-quality genetic maps were used to create a consensus linkage map involving 138,574 markers. This map was used to project the QTLs and conduct meta-QTL analysis. A total of 67 important meta-QTLs (MQTLs) were identified which were refined to 29 high-confidence MQTLs. The confidence interval (CI) of MQTLs ranged from 0 to 11.68 cM with a mean of 1.97 cM. The mean physical CI of MQTLs was 24.01 Mb, ranging from 0.0749 to 216.23 Mb per MQTL. As many as 44 MQTLs colocalized with marker-trait associations or SNP peaks associated with stripe rust resistance in wheat. Some MQTLs also included the following major genes- Yr5, Yr7, Yr16, Yr26, Yr30, Yr43, Yr44, Yr64, YrCH52, and YrH52. Candidate gene mining in high-confidence MQTLs identified 1,562 gene models. Examining these gene models for differential expressions yielded 123 differentially expressed genes, including the 59 most promising CGs. We also studied how these genes were expressed in wheat tissues at different phases of development. CONCLUSION: The most promising MQTLs identified in this study may facilitate marker-assisted breeding for stripe rust resistance in wheat. Information on markers flanking the MQTLs can be utilized in genomic selection models to increase the prediction accuracy for stripe rust resistance. The candidate genes identified can also be utilized for enhancing the wheat resistance against stripe rust after in vivo confirmation/validation using one or more of the following methods: gene cloning, reverse genetic methods, and omics approaches.

BACKGROUND: Bacterial epidemiology needs to understand the spread and dissemination of strains in a One Health context. This is important for highly pathogenic bacteria such as Bacillus anthracis, Brucella species, and Francisella tularensis. Whole genome sequencing (WGS) has paved the way for genetic marker detection and high-resolution genotyping. While such tasks are established for Illumina short-read sequencing, Oxford Nanopore Technology (ONT) long-read sequencing has yet to be evaluated for such highly pathogenic bacteria with little genomic variations between strains. In this study, three independent sequencing runs were performed using Illumina, ONT flow cell version 9.4.1, and 10.4 for six strains of each of Ba. anthracis, Br. suis and F. tularensis. Data from ONT sequencing alone, Illumina sequencing alone and two hybrid assembly approaches were compared. RESULTS: As previously shown, ONT produces ultra-long reads, while Illumina produces short reads with higher sequencing accuracy. Flow cell version 10.4 improved sequencing accuracy over version 9.4.1. The correct (sub-)species were inferred from all tested technologies, individually. Moreover, the sets of genetic markers for virulence, were almost identical for the respective species. The long reads of ONT allowed to assemble not only chromosomes of all species to near closure, but also virulence plasmids of Ba. anthracis. Assemblies based on nanopore data alone, Illumina data alone, and both hybrid assemblies correctly detected canonical (sub-)clades for Ba. anthracis and F. tularensis as well as multilocus sequence types for Br. suis. For F. tularensis, high-resolution genotyping using core-genome MLST (cgMLST) and core-genome Single-Nucleotide-Polymorphism (cgSNP) typing produced highly comparable results between data from Illumina and both ONT flow cell versions. For Ba. anthracis, only data from flow cell version 10.4 produced similar results to Illumina for both high-resolution typing methods. However, for Br. suis, high-resolution genotyping yielded larger differences comparing Illumina data to data from both ONT flow cell versions. CONCLUSIONS: In summary, combining data from ONT and Illumina for high-resolution genotyping might be feasible for F. tularensis and Ba. anthracis, but not yet for Br. suis. The ongoing improvement of nanopore technology and subsequent data analysis may facilitate high-resolution genotyping for all bacteria with highly stable genomes in future.

BACKGROUND: Genomic complexity is a growing field of evolution, with case studies for comparative evolutionary analyses in model and emerging non-model systems. Understanding complexity and the functional components of the genome is an untapped wealth of knowledge ripe for exploration. With the "remarkable lack of correspondence" between genome size and complexity, there needs to be a way to quantify complexity across organisms. In this study, we use a set of complexity metrics that allow for evaluating changes in complexity using TranD. RESULTS: We ascertain if complexity is increasing or decreasing across transcriptomes and at what structural level, as complexity varies. In this study, we define three metrics - TpG, EpT, and EpG- to quantify the transcriptome's complexity that encapsulates the dynamics of alternative splicing. Here we compare complexity metrics across 1) whole genome annotations, 2) a filtered subset of orthologs, and 3) novel genes to elucidate the impacts of orthologs and novel genes in transcript model analysis. Effective Exon Number (EEN) issued to compare the distribution of exon sizes within transcripts against random expectations of uniform exon placement. EEN accounts for differences in exon size, which is important because novel gene differences in complexity for orthologs and whole-transcriptome analyses are biased towards low-complexity genes with few exons and few alternative transcripts. CONCLUSIONS: With our metric analyses, we are able to quantify changes in complexity across diverse lineages with greater precision and accuracy than previous cross-species comparisons under ortholog conditioning. These analyses represent a step toward whole-transcriptome analysis in the emerging field of non-model evolutionary genomics, with key insights for evolutionary inference of complexity changes on deep timescales across the tree of life. We suggest a means to quantify biases generated in ortholog calling and correct complexity analysis for lineage-specific effects. With these metrics, we directly assay the quantitative properties of newly formed lineage-specific genes as they lower complexity.

BACKGROUND: Bread wheat is a recent allohexaploid (genomic constitution AABBDD) that emerged through a hybridization between tetraploid Triticum turgidum (AABB) and diploid Aegilops tauschii (DD) less than 10,000 years ago. The hexaploidization can be re-created artificially, producing synthetic wheat that has been used to study immediate genomic responses to polyploidization. The scale of the consequences of polyploidization, and their mechanism of establishment, remain uncertain. RESULTS: Here we sampled several synthetic wheats from alternative parental genotypes and reciprocal crosses, and examined transcriptomes from two different tissues and successive generations. We did not detect any massive reprogramming in gene expression, with only around 1% of expressed genes showing significant differences compared to their lower-ploidy parents. Most of this differential expression is located on the D subgenome, without consistency in the direction of the expression change. Homoeolog expression bias in synthetic wheat is similar to the pattern observed in the parents. Both differential expression and homoeolog bias are tissue-specific. While up to three families of transposable elements became upregulated in wheat synthetics, their position and distance are not significantly associated with expression changes in proximal genes. DISCUSSION: While only a few genes change their expression pattern after polyploidization, they can be involved in agronomically important pathways. Alternative parental combinations can lead to opposite changes on the same subset of D-located genes, which is relevant for harnessing new diversity in wheat breeding. Tissue specificity of the polyploidization-triggered expression changes indicates the remodelling of transcriptomes in synthetic wheat is plastic and likely caused by regulome interactions rather than permanent changes. We discuss the pitfalls of transcriptomic comparisons across ploidy levels that can inflate the de-regulation signal. CONCLUSIONS: Transcriptomic response to polyploidization in synthetic AABBDD wheat is modest and much lower than some previous estimates. Homoeolog expression bias in wheat allohexaploids is mostly attributed to parental legacy, with polyploidy having a mild balancing effect.

BACKGROUND: As an important source of genetic variation, copy number variation (CNV) can alter the dosage of DNA segments, which in turn may affect gene expression level and phenotype. However, our knowledge of CNV in apple is still limited. Here, we obtained high-confidence CNVs and investigated their functional impact based on genome resequencing data of two apple populations, cultivars and wild relatives. RESULTS: In this study, we identified 914,610 CNVs comprising 14,839 CNV regions (CNVRs) from 346 apple accessions, including 289 cultivars and 57 wild relatives. CNVRs summed to 71.19 Mb, accounting for 10.03% of the apple genome. Under the low linkage disequilibrium (LD) with nearby SNPs, they could also accurately reflect the population structure of apple independent of SNPs. Furthermore, A total of 3,621 genes were covered by CNVRs and functionally involved in biological processes such as defense response, reproduction and metabolic processes. In addition, the population differentiation index ([Formula: see text]) analysis between cultivars and wild relatives revealed 127 CN-differentiated genes, which may contribute to trait differences in these two populations. CONCLUSIONS: This study was based on identification of CNVs from 346 diverse apple accessions, which to our knowledge was the largest dataset for CNV analysis in apple. Our work presented the first comprehensive CNV map and provided valuable resources for understanding genomic variations in apple.

Cis-regulatory elements (CRE) are critical for coordinating gene expression programs that dictate cell-specific differentiation and homeostasis. Recently developed self-transcribing active regulatory region sequencing (STARR-Seq) has allowed for genome-wide annotation of functional CREs. Despite this, STARR-Seq assays are only employed in cell lines, in part, due to difficulties in delivering reporter constructs. Herein, we implemented and validated a STARR-Seq-based screen in human CD4+ T cells using a non-integrating lentiviral transduction system. Lenti-STARR-Seq is the first example of a genome-wide assay of CRE function in human primary cells, identifying thousands of functional enhancers and negative regulatory elements (NREs) in human CD4+ T cells. We find an unexpected difference in nucleosome organization between enhancers and NRE: enhancers are located between nucleosomes, whereas NRE are occupied by nucleosomes in their endogenous locations. We also describe chromatin modification, eRNA production, and transcription factor binding at both enhancers and NREs. Our findings support the idea of silencer repurposing as enhancers in alternate cell types. Collectively, these data suggest that Lenti-STARR-Seq is a successful approach for CRE screening in primary human cell types, and provides an atlas of functional CREs in human CD4+ T cells.

BACKGROUND: The long non-coding RNAs (lncRNAs) are critical regulators of diverse biological processes. Nevertheless, a global view of its expression and function in the mouse retina, a crucial model for neurogenesis study, still needs to be made available. RESULTS: Herein, by integrating the established gene models and the result from ab initio prediction using short- and long-read sequencing, we characterized 4,523 lncRNA genes (MRLGs) in developing mouse retinas (from the embryonic day of 12.5 to the neonatal day of P28), which was so far the most comprehensive collection of retinal lncRNAs. Next, derived from transcriptomics analyses of different tissues and developing retinas, we found that the MRLGs were highly spatiotemporal specific in expression and played essential roles in regulating the genesis and function of mouse retinas. In addition, we investigated the expression of MRLGs in some mouse mutants and revealed that 97 intergenic MRLGs might be involved in regulating differentiation and development of retinal neurons through Math5, Isl1, Brn3b, NRL, Onecut1, or Onecut2 mediated pathways. CONCLUSIONS: In summary, this work significantly enhanced our knowledge of lncRNA genes in mouse retina development and provided valuable clues for future exploration of their biological roles.

With the rising demand for entry to extremely high altitudes (HAs), rapid adaptability to extremely hypoxic environments is a challenge that we need to explore. Fasting was used to evaluate acute hypoxia tolerance at HA and was proven to be an effective method for improving the survival rate at extreme HA. Our experiments also showed that fasting pretreatment for 72 h significantly increased the 24 h survival rate of rats at 7620 m from 10 to 85% and protected the myocardium cells of rats. Here, we compared the metabolites and gene expression in the myocardium of SD rats pretreated with fasting and nonfasting at normal altitude and extreme HA. Our findings demonstrated that the dynamic contents of detected differential metabolites (DMs) between different rat groups were consistent with the expression of differentially expressed genes (DEGs), and DM clusters also showed strong correlations with DEG clusters. DM clusters related to amino acids and lipids were significantly lower in the fasting groups, and the correlated DEG clusters were enriched in mitotic pathways, including CDK1, CDC7, NUF2, and MCM6, suggesting that fasting can attenuate mitotic processes in cardiac tissues and reduce the synthesis of amino acids and lipids. L-Glutamine-related metabolites were particularly low at extreme HA without pretreatment but were normal in the fasting groups. The DEGs in the cluster related to L-glutamine-related metabolites were enriched for T-cell receptor V(D)J recombination, the Hippo signaling pathway, the Wnt signaling pathway, the cGMP-PKG signaling pathway, and the mTOR signaling pathway and were significantly downregulated, indicating that the content of L-glutamine decreased at extreme HA, while fasting increased it to adapt to the environment. Moreover, abundant fatty acids were detected when rats were exposed to extreme HA without pretreatment. Our study revealed the fasting and hypoxic environment-related factors in SD rats and provided new insights into the genetic and molecular characteristics in the myocardium, which is critical to developing more potential rapid adaptation methods to extreme HA.

BACKGROUND: Real-time quantitative PCR (RT-qPCR) is a crucial and widely used method for gene expression analysis. Selecting suitable reference genes is extremely important for the accuracy of RT-qPCR results. Commonly used reference genes are not always stable in various organisms or under different environmental conditions. With the increasing application of high-throughput sequencing, transcriptome analysis has become an effective method for identifying novel stable reference genes. RESULTS: In this study, we identified candidate reference genes based on transcriptome data covering embryos and larvae of early development, normal adult tissues, and the hindgut under sulfide stress using the coefficient of variation (CV) method in the echiuran Urechis unicinctus, resulting in 6834 (15.82%), 7110 (16.85%) and 13880 (35.87%) candidate reference genes, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that the candidate reference genes were significantly enriched in cellular metabolic process, protein metabolic process and ribosome in early development and normal adult tissues as well as in cellular localization and endocytosis in the hindgut under sulfide stress. Subsequently, ten genes including five new candidate reference genes and five commonly used reference genes, were validated by RT-qPCR. The expression stability of the ten genes was analyzed using four methods (geNorm, NormFinder, BestKeeper, and ∆Ct). The comprehensive results indicated that the new candidate reference genes were more stable than most commonly used reference genes. The commonly used ACTB was the most unstable gene. The candidate reference genes STX12, EHMT1, and LYAG were the most stable genes in early development, normal adult tissues, and hindgut under sulfide stress, respectively. The log2(TPM) of the transcriptome data was significantly negatively correlated with the Ct values of RT-qPCR (Ct =  - 0.5405 log2(TPM) + 34.51), which made it possible to estimate the Ct value before RT-qPCR using transcriptome data. CONCLUSION: Our study is the first to select reference genes for RT-qPCR from transcriptome data in Echiura and provides important information for future gene expression studies in U. unicinctus.

BACKGROUND: Marine deep subsurface sediments were once thought to be devoid of eukaryotic life, but advances in molecular technology have unlocked the presence and activity of well-known closely related terrestrial and marine fungi. Commonly detected fungi in deep marine sediment environments includes Penicillium, Aspergillus, Cladosporium, Fusarium, and Schizophyllum, which could have important implications in carbon and nitrogen cycling in this isolated environment. In order to determine the diversity and unknown metabolic capabilities of fungi in deep-sea sediments, their genomes need to be fully analyzed. In this study, two Penicillium species were isolated from South Pacific Gyre sediment enrichments during Integrated Ocean Drilling Program Expedition 329. The inner gyre has very limited productivity, organic carbon, and nutrients. RESULTS: Here, we present high-quality genomes of two proposed novel Penicillium species using Illumina HiSeq and PacBio sequencing technologies. Single-copy homologues within the genomes were compared to other closely related genomes using OrthoMCL and maximum-likelihood estimation, which showed that these genomes were novel species within the genus Penicillium. We propose to name isolate SPG-F1 as Penicillium pacificasedimenti sp. nov. and SPG-F15 as Penicillium pacificagyrus sp. nov. The resulting genome sizes were 32.6 Mbp and 36.4 Mbp, respectively, and both genomes were greater than 98% complete as determined by the presence of complete single-copy orthologs. The transposable elements for each genome were 4.87% for P. pacificasedimenti and 10.68% for P. pacificagyrus. A total of 12,271 genes were predicted in the P. pacificasedimenti genome and 12,568 genes in P. pacificagyrus. Both isolates contained genes known to be involved in the degradation of recalcitrant carbon, amino acids, and lignin-derived carbon. CONCLUSIONS: Our results provide the first constructed genomes of novel Penicillium isolates from deep marine sediments, which will be useful for future studies of marine subsurface fungal diversity and function. Furthermore, these genomes shed light on the potential impact fungi in marine sediments and the subseafloor could have on global carbon and nitrogen biogeochemical cycles and how they may be persisting in the most energy-limited sedimentary biosphere.

BACKGROUND: Rootstocks can improve the chilling tolerance of grafted cucumbers, but their effectiveness varies. Rootstocks with strong de-blooming capacity may result in lower chilling tolerance of grafted cucumbers compared to those with weak de-blooming capacity, while also reducing the silicon absorption. However, it remains unclear whether this reduction in chilling tolerance is due to differences in rootstock genotypes or the reduction in silicon absorption. RESULTS: The chilling tolerance of cucumber seedlings was improved by using rootstocks and silicon nutrition. Rootstocks had a more significant effect than silicon nutrition, and the weak de-blooming rootstock 'Yunnan figleaf gourd' was superior to the strong de-blooming rootstock 'Huangchenggen No. 2'. Compared to self-rooted cucumber, twelve miRNAs were regulated by two rootstocks, including seven identical miRNAs (novel-mir23, novel-mir26, novel-mir30, novel-mir37, novel-mir46, miR395a and miR398a-3p) and five different miRNAs (novel-mir32, novel-mir38, novel-mir65, novel-mir78 and miR397a). Notably, four of these miRNAs (novel-mir38, novel-mir65, novel-mir78 and miR397a) were only identified in 'Yunnan figleaf gourd'-grafted cucumbers. Furthermore, six miRNAs (miR168a-5p, miR390a-5p, novel-mir26, novel-mir55, novel-mir67 and novel-mir70) were found to be responsive to exogenous silicon. Target gene prediction for 20 miRNAs resulted in 520 genes. Functional analysis of these target genes showed that 'Yunnan figleaf gourd' improves the chilling tolerance of cucumber by regulating laccase synthesis and sulfate metabolism, while 'Huangchenggen No. 2' and exogenous silicon reduced chilling stress damage to cucumber by regulating ROS scavenging and protein protection, respectively. CONCLUSION: Among the identified miRNAs, novel-mir46 and miR398a-3p were found in cucumbers in response to chilling stress and two types of rootstocks. However, no identical miRNAs were identified in response to chilling stress and silicon. In addition, the differential expression of novel-mir38, novel-mir65, novel-mir78 and miR397a may be one of the important reasons for the differences in chilling tolerance of grafted cucumbers caused by two types of rootstocks.

Open reading frames (ORFs) with fewer than 100 codons are generally not annotated in genomes, although bona fide genes of that size are known. Newer biochemical studies have suggested that thousands of small protein-coding ORFs (smORFs) may exist in the human genome, but the true number and the biological significance of the micropeptides they encode remain uncertain. Here, we used a comparative genomics approach to identify high-confidence smORFs that are likely protein-coding. We identified 3,326 high-confidence smORFs using constraint within human populations and evolutionary conservation as additional lines of evidence. Next, we validated that, as a group, our high-confidence smORFs are conserved at the amino-acid level rather than merely residing in highly conserved non-coding regions. Finally, we found that high-confidence smORFs are enriched among disease-associated variants from GWAS. Overall, our results highlight that smORF-encoded peptides likely have important functional roles in human disease.

BACKGROUND: Structural variants (SVs) are chromosomal segments that differ between genomes, such as deletions, duplications, insertions, inversions and translocations. The genomics revolution enabled the discovery of sub-microscopic SVs via array and whole-genome sequencing (WGS) data, paving the way to unravel the functional impact of SVs. Recent human expression QTL mapping studies demonstrated that SVs play a disproportionally large role in altering gene expression, underlining the importance of including SVs in genetic analyses. Therefore, this study aimed to generate and explore a high-quality bovine SV catalogue exploiting a unique cattle family cohort data (total 266 samples, forming 127 trios). RESULTS: We curated 13,731 SVs segregating in the population, consisting of 12,201 deletions, 1,509 duplications, and 21 multi-allelic CNVs (> 50-bp). Of these, we validated a subset of copy number variants (CNVs) utilising a direct genotyping approach in an independent cohort, indicating that at least 62% of the CNVs are true variants, segregating in the population. Among gene-disrupting SVs, we prioritised two likely high impact duplications, encompassing ORM1 and POPDC3 genes, respectively. Liver expression QTL mapping results revealed that these duplications are likely causing altered gene expression, confirming the functional importance of SVs. Although most of the accurately genotyped CNVs are tagged by single nucleotide polymorphisms (SNPs) ascertained in WGS data, most CNVs were not captured by individual SNPs obtained from a 50K genotyping array. CONCLUSION: We generated a high-quality SV catalogue exploiting unique whole genome sequenced bovine family cohort data. Two high impact duplications upregulating the ORM1 and POPDC3 are putative candidates for postpartum feed intake and hoof health traits, thus warranting further investigation. Generally, CNVs were in low LD with SNPs on the 50K array. Hence, it remains crucial to incorporate CNVs via means other than tagging SNPs, such as investigation of tagging haplotypes, direct imputation of CNVs, or direct genotyping as done in the current study. The SV catalogue and the custom genotyping array generated in the current study will serve as valuable resources accelerating utilisation of full spectrum of genetic variants in bovine genomes.

BACKGROUND: The receptor-like kinase (RLK) gene families in plants contains a large number of members. They are membrane proteins with an extracellular receptor domain and participate in biotic and abiotic stress responses. RESULTS: In this study, we identified RLKs in 15 representative plant genomes, including wheat, and classified them into 64 subfamilies by using four types of phylogenetic trees and HMM models. Conserved exon‒intron structures with conserved exon phases in the kinase domain were found in many RLK subfamilies from Physcomitrella patens to Triticum aestivum. Domain distributions of RLKs were also diagrammed. Collinearity events and tandem gene clusters suggested that polyploidization and tandem duplication events contributed to the member expansions of T. aestivum RLKs. Global expression pattern analysis was performed by using public transcriptome data. These analyses were involved in T. aestivum, Aegilops tauschii and Brachypodium distachyon RLKs under biotic and abiotic stresses. We also selected 9 RLKs to validate the transcriptome prediction by using qRT‒PCR under drought treatment and with Fusarium graminearum infection. The expression trends of these 9 wheat RLKs from public transcriptome data were consistent with the results of qRT‒PCR, indicating that they might be stress response genes under drought or F. graminearum treatments. CONCLUSION: In this study, we identified, classified, evolved, and expressed RLKs in wheat and related plants. Thus, our results will provide insights into the evolutionary history and molecular mechanisms of wheat RLKs.