The 1000 Genomes Project (1kGP) is the largest fully open resource of whole-genome sequencing (WGS) data consented for public distribution without access or use restrictions. The final, phase 3 ...release of the 1kGP included 2,504 unrelated samples from 26 populations and was based primarily on low-coverage WGS. Here, we present a high-coverage 3,202-sample WGS 1kGP resource, which now includes 602 complete trios, sequenced to a depth of 30X using Illumina. We performed single-nucleotide variant (SNV) and short insertion and deletion (INDEL) discovery and generated a comprehensive set of structural variants (SVs) by integrating multiple analytic methods through a machine learning model. We show gains in sensitivity and precision of variant calls compared to phase 3, especially among rare SNVs as well as INDELs and SVs spanning frequency spectrum. We also generated an improved reference imputation panel, making variants discovered here accessible for association studies.
The Pacific oyster Crassostrea gigas belongs to one of the most species-rich but genomically poorly explored phyla, the Mollusca. Here we report the sequencing and assembly of the oyster genome using ...short reads and a fosmid-pooling strategy, along with transcriptomes of development and stress response and the proteome of the shell. The oyster genome is highly polymorphic and rich in repetitive sequences, with some transposable elements still actively shaping variation. Transcriptome studies reveal an extensive set of genes responding to environmental stress. The expansion of genes coding for heat shock protein 70 and inhibitors of apoptosis is probably central to the oyster's adaptation to sessile life in the highly stressful intertidal zone. Our analyses also show that shell formation in molluscs is more complex than currently understood and involves extensive participation of cells and their exosomes. The oyster genome sequence fills a void in our understanding of the Lophotrochozoa.
The Pacific oyster, Crassostrea gigas, has developed special mechanisms to regulate its osmotic balance to adapt to fluctuations of salinities in coastal zones. To understand the oyster's euryhaline ...adaptation, we analyzed salt stress effectors metabolism pathways under different salinities (salt 5, 10, 15, 20, 25, 30 and 40 for 7 days) using transcriptome data, physiology experiment and quantitative real-time PCR.
Transcriptome data uncovered 189, 480, 207 and 80 marker genes for monitoring physiology status of oysters and the environment conditions. Three known salt stress effectors (involving ion channels, aquaporins and free amino acids) were examined. The analysis of ion channels and aquaporins indicated that 7 days long-term salt stress inhibited voltage-gated Na(+)/K(+) channel and aquaporin but increased calcium-activated K(+) channel and Ca(2+) channel. As the most important category of osmotic stress effector, we analyzed the oyster FAAs metabolism pathways (including taurine, glycine, alanine, beta-alanine, proline and arginine) and explained FAAs functional mechanism for oyster low salinity adaptation. FAAs metabolism key enzyme genes displayed expression differentiation in low salinity adapted individuals comparing with control which further indicated that FAAs played important roles for oyster salinity adaptation. A global metabolic pathway analysis (iPath) of oyster expanded genes displayed a co-expansion of FAAs metabolism in C. gigas compared with seven other species, suggesting oyster's powerful ability regarding FAAs metabolism, allowing it to adapt to fluctuating salinities, which may be one important mechanism underlying euryhaline adaption in oyster. Additionally, using transcriptome data analysis, we uncovered salt stress transduction networks in C. gigas.
Our results represented oyster salt stress effectors functional mechanisms under salt stress conditions and explained the expansion of FAAs metabolism pathways as the most important effectors for oyster euryhaline adaptation. This study was the first to explain oyster euryhaline adaptation at a genome-wide scale in C. gigas.
• The A-genome group in Oryza consists of eight diploid species and is distributed world-wide. Here we reconstructed the phylogeny among the A-genome species based on sequences of nuclear genes and ...MITE (miniature inverted-repeat transposable elements) insertions. • Thirty-seven accessions representing two cultivated and six wild species from the A-genome group were sampled. Introns of four nuclear single-copy genes on different chromosomes were sequenced and analysed by both maximum parsimony (MP) and Bayesian inference methods. • All the species except for Oryza rufipogon and Oryza nivara formed a monophyletic group and the Australian endemic Oryza meridionalis was the earliest divergent lineage. Two subspecies of Oryza sativa (ssp. indica and ssp. japonica) formed two separate monophyletic groups, suggestive of their polyphyletic origin. Based on molecular clock approach, we estimated that the divergence of the A-genome group occurred c. 2.0 million years ago (mya) while the two subspecies (indica and japonica) separated c. 0.4 mya. • Intron sequences of nuclear genes provide sufficient resolution and are informative for phylogenetic inference at lower taxonomic levels.
The genus Liriodendron belongs to the family Magnoliaceae, which resides within the magnoliids, an early diverging lineage of the Mesangiospermae. However, the phylogenetic relationship of magnoliids ...with eudicots and monocots has not been conclusively resolved and thus remains to be determined
. Liriodendron is a relict lineage from the Tertiary with two distinct species-one East Asian (L. chinense (Hemsley) Sargent) and one eastern North American (L. tulipifera Linn)-identified as a vicariad species pair. However, the genetic divergence and evolutionary trajectories of these species remain to be elucidated at the whole-genome level
. Here, we report the first de novo genome assembly of a plant in the Magnoliaceae, L. chinense. Phylogenetic analyses suggest that magnoliids are sister to the clade consisting of eudicots and monocots, with rapid diversification occurring in the common ancestor of these three lineages. Analyses of population genetic structure indicate that L. chinense has diverged into two lineages-the eastern and western groups-in China. While L. tulipifera in North America is genetically positioned between the two L. chinense groups, it is closer to the eastern group. This result is consistent with phenotypic observations that suggest that the eastern and western groups of China may have diverged long ago, possibly before the intercontinental differentiation between L. chinense and L. tulipifera. Genetic diversity analyses show that L. chinense has tenfold higher genetic diversity than L. tulipifera, suggesting that the complicated regions comprising east-west-orientated mountains and the Yangtze river basin (especially near 30° N latitude) in East Asia offered more successful refugia than the south-north-orientated mountain valleys in eastern North America during the Quaternary glacial period.
The Pacific oyster
Crassostrea gigas
, a commercially important species inhabiting the intertidal zone, facing enormous temperature fluctuations. Therefore, it is important to identify candidate ...genes and key regulatory relationships associated with thermal tolerance, which can aid the molecular breeding of oysters. Heat shock transcription factor 1 (
HSF1
) plays an important role in the thermal stress resistance. However, the regulatory relationship between the expansion of heat shock protein (HSP)
HSP
70 and HSF1 is not yet clear in
C. gigas
. In this study, we analyzed genes regulated by
HSF1
in response to heat shock by chromatin immunoprecipitation followed by sequencing (ChIP-seq), determined the expression patterns of target genes by qRT-PCR, and validated the regulatory relationship between one
HSP70
and HSF1. We found 916 peaks corresponding to HSF1 binding sites, and these peaks were annotated to the nearest genes. In Gene Ontology analysis, HSF1 target genes were related to signal transduction, energy production, and response to biotic stimulus. Four
HSP70
genes, two
HSP40
genes, and one small
HSP
gene exhibited binding to HSF1. One
HSP70
with a binding site in the promoter region was validated to be regulated by HSF1 under heat shock. These results provide a basis for future studies aimed at determining the mechanisms underlying thermal tolerance and provide insights into gene regulation in the Pacific oyster.
Varying degrees of reduction of genetic diversity in crops relative to their wild progenitors occurred during the process of domestication. Such information, however, has not been available for the ...Asian cultivated rice (Oryza sativa) despite its importance as a staple food and a model organism. To reveal levels and patterns of nucleotide diversity and to elucidate the genetic relationship and demographic history of O. sativa and its close relatives (Oryza rufipogon and Oryza nivara), we investigated nucleotide diversity data from 10 unlinked nuclear loci in species-wide samples of these species. The results indicated that O. rufipogon and O. nivara possessed comparable levels of nucleotide variation (θsil = 0.0077∼0.0095) compared with the relatives of other crops. In contrast, nucleotide diversity of O. sativa was as low as θsil = 0.0024 and even lower (θsil = 0.0021 for indica and 0.0011 for japonica), if we consider the 2 subspecies separately. Overall, only 20-10% of the diversity in the wild species was retained in 2 subspecies of the cultivated rice (indica and japonica), respectively. Because statistic tests did not reject the assumption of neutrality for all 10 loci, we further used coalescent to simulate bottlenecks under various lengths and population sizes to better understand the domestication process. Consistent with the dramatic reduction in nucleotide diversity, we detected a severe domestication bottleneck and demonstrated that the sequence diversity currently found in the rice genome could be explained by a founding population of 1,500 individuals if the initial domestication event occurred over a 3,000-year period. Phylogenetic analyses revealed close genetic relationships and ambiguous species boundary of O. rufipogon and O. nivara, providing additional evidence to treat them as 2 ecotypes of a single species. Lowest linkage disequilibrium (LD) was found in the perennial O. rufipogon where the r
2 value dropped to a negligible level within 400 bp, and the highest in the japonica rice where LD extended to the entirely sequenced region (∼900 bp), implying that LD mapping by genome scans may not be feasible in wild rice due to the high density of markers needed.
Rock bream (
Oplegnathus fasciatus
) is a valuable commercial marine teleost species, which exhibits sexual dimorphism in growth performance. However, the absence of a rapid and cost-effective sex ...identification method based on sex-specific genetic marker has impeded study on sex determination mechanisms and breeding applications. In the present study, we firstly developed the PCR method for identifying potential sex-specific sequences in
Oplegnathus fasciatus
with the next-generation sequencing. Sex-specific genomic regions/loci for sex determination were discovered on Chr2 and Chr6 by genome-wide association analysis, sequencing depth, and heterozygosity comparison between females and males. Candidate sex-determining genes (
CCDC63
,
ITR
,
WNT4
) were furtherly detected in transcriptome data of testes and ovaries. Taken together, a male-specific 34-bp deletion on the Chr2 was identified and developed into molecular marker of sex for
O
.
fasciatus
. After validation in individuals with known phenotypic sexes, the accuracy was 100%. This study gives an insight into the mechanism of sex determination in
O
.
fasciatus
, and the gender marker is crucial both for future genomic research and for development of efficient and sustainable aquaculture practice.
The rock bream (
) is a typical fish with a unique multiple sex chromosome system. In this study, we investigated the gene expression profiling in the gonads and brains of both males and females ...using RNA-Seq to identify sex-related genes and pathways. In accordance with the dimorphic expression profiles, combined with Gene ontology and KEGG enrichment analyses, a number of potential genes and pathways associated with sex determination were obtained from transcriptional analysis, especially some sex-biased genes and pathways. Next, we selected 18 candidate genes and analyzed their expression in different tissues and developmental stages. We found that the expression levels of
,
,
,
, and
were significantly higher in the testis than those in the ovary or other tissues, whereas the expression levels of
,
,
,
, and
were significantly higher in the ovary than those in the testis. Furthermore, the expression levels of these genes in different developmental stages of gonads also showed sexually dimorphic patterns, suggesting that they might play important roles during gonadal development. These genes are useful markers for investigating sex determination and differentiation in rock bream. The findings of this study can provide insights into the molecular mechanisms of sex determination and differentiation in fish with multiple sex chromosome systems.
The persistence of covalently closed circular DNA (cccDNA) in infected hepatocytes is the major barrier preventing viral eradication with existing therapies in patients with chronic hepatitis B. ...Therapeutic agents that can eliminate cccDNA are urgently needed to achieve viral eradication and thus HBV cure.
A phenotypic assay with HBV-infected primary human hepatocytes (PHHs) was employed to screen for novel cccDNA inhibitors. A HBVcircle mouse model and a uPA-SCID (urokinase-type plasminogen activator-severe combined immunodeficiency) humanized liver mouse model were used to evaluate the anti-HBV efficacy of the discovered cccDNA inhibitors.
Potent and dose-dependent reductions in extracellular HBV DNA, HBsAg, and HBeAg levels were achieved upon the initiation of ccc_R08 treatment two days after the HBV infection of PHHs. More importantly, the level of cccDNA was specifically reduced by ccc_R08, while it did not obviously affect mitochondrial DNA. Additionally, ccc_R08 showed no significant cytotoxicity in PHHs or in multiple proliferating cell lines. The twice daily oral administration of ccc_R08 to HBVcircle model mice, which contained surrogate cccDNA molecules, significantly decreased the serum levels of HBV DNA and antigens, and these effects were sustained during the off-treatment follow-up period. Moreover, at the end of follow-up, the levels of surrogate cccDNA molecules in the livers of ccc_R08-treated HBVcircle mice were reduced to below the lower limit of quantification.
We have discovered a small-molecule cccDNA inhibitor that reduces HBV cccDNA levels. cccDNA inhibitors potentially represent a new approach to completely cure patients chronically infected with HBV.
Covalently closed circular DNA (cccDNA) persistence in HBV-infected hepatocytes is the root cause of chronic hepatitis B. We discovered a novel small-molecule cccDNA inhibitor that can specifically reduce cccDNA levels in HBV-infected hepatocytes. This type of molecule could offer a new approach to completely cure patients chronically infected with HBV.
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•A robust in vitro primary human hepatocyte HBV infection assay protocol is discovered for screening cccDNA inhibitors.•ccc_R08 is a novel orally available HBV cccDNA inhibitor that could reduce the level of cccDNA in infected primary human hepatocytes.•ccc_R08 treatment leads to sustained HBsAg and cccDNA level reductions in the HBVcircle mouse model.•pgRNA reduction quantitatively correlates with liver cccDNA reduction.
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