The diploid genome sequence of an Asian individual

Here we present the first diploid genome sequence of an Asian individual. The genome was sequenced to 36-fold average coverage using massively parallel sequencing technology. We aligned the short reads onto the NCBI human reference genome to 99.97% coverage, and guided by the reference genome, we used uniquely mapped reads to assemble a high-quality consensus sequence for 92% of the Asian individual's genome. We identified approximately 3 million single-nucleotide polymorphisms (SNPs) inside this region, of which 13.6% were not in the dbSNP database. Genotyping analysis showed that SNP identification had high accuracy and consistency, indicating the high sequence quality of this assembly. We also carried out heterozygote phasing and haplotype prediction against HapMap CHB and JPT haplotypes (Chinese and Japanese, respectively), sequence comparison with the two available individual genomes (J. D. Watson and J. C. Venter), and structural variation identification. These variations were considered for their potential biological impact. Our sequence data and analyses demonstrate the potential usefulness of next-generation sequencing technologies for personal genomics.     

Sequence identification of 2,375 human brain genes.

We recently described a new approach for the rapid characterization of expressed genes by partial DNA sequencing to generate 'expressed sequence tags'. From a set of 600 human brain complementary DNA clones, 348 were informative nuclear-encoded messenger RNAs. We have now partially sequenced 2,672 new, independent cDNA clones isolated from four human brain cDNA libraries to generate 2,375 expressed sequence tags to nuclear-encoded genes. These sequences, together with 348 brain expressed sequence tags from our previous study, comprise more than 2,500 new human genes and 870,769 base pairs of DNA sequence. These data represent an approximate doubling of the number of human genes identified by DNA sequencing and may represent as many as 5% of the genes in the human genome.     

利用人类全基因组二代测序数据比较BWA-MEM 和 NovoAlign

随着测序技术的发展,二代测序数据越来越多,将测序数据准确地比对到参考基因组是后续研究的基础．BWA-MEM和NovoAlign作为2个最常用的DNA序列比对软件,还没有评估其在基因组中不同结构区域的表现．本研究基于真实和模拟数据,对2个软件在人类基因组的低复杂度、片段性重复和其他区域进行了评估．结果显示:BWA-MEM将尽可能多的测序数据比对到基因组,且在低复杂度和片段性重复区域存在过度比对的现象,特别是在片段性重复区域的比对品质较低;而NovoAlign比对到基因组的序列数量低于BWA-MEM,但在片段性重复区域的比对品质较优,因此对于存在较多片段性重复区域的基因组来说,使用NovoAlign比对是一种更好的策略．      

An SNP map of human chromosome 22

The human genome sequence will provide a reference for measuring DNA sequence variation in human populations. Sequence variants are responsible for the genetic component of individuality, including complex characteristics such as disease susceptibility and drug response. Most sequence variants are single nucleotide polymorphisms (SNPs), where two alternate bases occur at one position. Comparison of any two genomes reveals around 1 SNP per kilobase. A sufficiently dense map of SNPs would allow the detection of sequence variants responsible for particular characteristics on the basis that they are associated with a specific SNP allele. Here we have evaluated large-scale sequencing approaches to obtaining SNPs, and have constructed a map of 2,730 SNPs on human chromosome 22. Most of the SNPs are within 25 kilobases of a transcribed exon, and are valuable for association studies. We have scaled up the process, detecting over 65,000 SNPs in the genome as part of The SNP Consortium programme, which is on target to build a map of 1 SNP every 5 kilobases that is integrated with the human genome sequence and that is freely available in the public domain.     

林木分子育种研究的基因组学信息资源述评

分子育种是指利用与性状相关的DNA标记进行选育,也称标记辅助选择或标记辅助育种,广义上还包括基因工程育种和基因组学辅助育种。林木分子育种为早期选择和加速育种提供了极具潜力的高效手段。笔者对林木分子育种研究的基因组学信息资源进行了进展综述和前景展望。近30年来,林木分子标记技术从早期的低通量方法发展到目前基于微阵列芯片和新一代测序的高通量技术,如测序分型、转录组测序、重测序、扩增子测序和外显子组测序等,并广泛用于连锁作图、关联分析和基因组选择等林木性状相关的DNA变异检测研究。随着2006年毛果杨基因组序列的发表,已有50余个树种完成了基因组测序。基于连锁作图和关联研究检测了林木10余个属生长、材性和抗逆及非木质产品品质等性状相关的大量基因组位点,主要趋势表现为：① 表型广泛,涵盖经济性状、生理指标和代谢成分等;②标记数量成千上万甚至上百万,覆盖全基因组;③转录组和降解组等多组学的分子变异开始应用;④ 利用大群体以提高位点检测的精度;⑤ 重视环境的影响,大田试验设置多个地点,解析QTL与环境、年份的互作效应;⑥ 结合参考基因组序列和/或转录组差异表达基因进一步挖掘性状相关的候选基因,建立了桉属、松属和云杉属等主要造林树种的基因组选择模型。此外,积累了泛基因组、相关软件和算法、功能基因、基因组编辑技术及网站和数据库等其他信息资源。林木分子育种面临的挑战主要包括：① 如何获得稳定性好的性状相关基因组位点和基因组选择（GS）模型;② 缺乏自动化、无损和高通量的表型测定技术;③对大基因组的针叶树和一些多倍体树种,仍难获得高质量的基因组序列;④ 标记辅助选择增加了常规育种之外的费用,且存在不确定性;⑤多数树种的加速育种仍较困难。后基因组时代的林木分子育种将有效结合到常规育种程序中,显著促进遗传增益的提高。     

The DNA sequence and biological annotation of human chromosome 1

The reference sequence for each human chromosome provides the framework for understanding genome function, variation and evolution. Here we report the finished sequence and biological annotation of human chromosome 1. Chromosome 1 is gene-dense, with 3,141 genes and 991 pseudogenes, and many coding sequences overlap. Rearrangements and mutations of chromosome 1 are prevalent in cancer and many other diseases. Patterns of sequence variation reveal signals of recent selection in specific genes that may contribute to human fitness, and also in regions where no function is evident. Fine-scale recombination occurs in hotspots of varying intensity along the sequence, and is enriched near genes. These and other studies of human biology and disease encoded within chromosome 1 are made possible with the highly accurate annotated sequence, as part of the completed set of chromosome sequences that comprise the reference human genome.     

The complete genome of an individual by massively parallel DNA sequencing

The association of genetic variation with disease and drug response, and improvements in nucleic acid technologies, have given great optimism for the impact of 'genomic medicine'. However, the formidable size of the diploid human genome, approximately 6 gigabases, has prevented the routine application of sequencing methods to deciphering complete individual human genomes. To realize the full potential of genomics for human health, this limitation must be overcome. Here we report the DNA sequence of a diploid genome of a single individual, James D. Watson, sequenced to 7.4-fold redundancy in two months using massively parallel sequencing in picolitre-size reaction vessels. This sequence was completed in two months at approximately one-hundredth of the cost of traditional capillary electrophoresis methods. Comparison of the sequence to the reference genome led to the identification of 3.3 million single nucleotide polymorphisms, of which 10,654 cause amino-acid substitution within the coding sequence. In addition, we accurately identified small-scale (2-40,000 base pair (bp)) insertion and deletion polymorphism as well as copy number variation resulting in the large-scale gain and loss of chromosomal segments ranging from 26,000 to 1.5 million base pairs. Overall, these results agree well with recent results of sequencing of a single individual by traditional methods. However, in addition to being faster and significantly less expensive, this sequencing technology avoids the arbitrary loss of genomic sequences inherent in random shotgun sequencing by bacterial cloning because it amplifies DNA in a cell-free system. As a result, we further demonstrate the acquisition of novel human sequence, including novel genes not previously identified by traditional genomic sequencing. This is the first genome sequenced by next-generation technologies. Therefore it is a pilot for the future challenges of 'personalized genome sequencing'.     

A physical map of the human genome

The human genome is by far the largest genome to be sequenced, and its size and complexity present many challenges for sequence assembly. The International Human Genome Sequencing Consortium constructed a map of the whole genome to enable the selection of clones for sequencing and for the accurate assembly of the genome sequence. Here we report the construction of the whole-genome bacterial artificial chromosome (BAC) map and its integration with previous landmark maps and information from mapping efforts focused on specific chromosomal regions. We also describe the integration of sequence data with the map.     

全基因组测序技术研究及其在木本植物中的应用

基因组序列是开展遗传研究重要的信息基础,随着测序技术飞速发展至第3代长片段测序方法,测序读长历经从几十到数万个碱基的提升,对进一步提升基因组组装的完整度以及准确性提供了极大的裨益。现已完成了大量植物种全基因组测序工作,其中木本植物有40多个,还有更多树种的全基因组测序正在进行之中。针对各类测序技术的基因组组装及后续分析,研究人员也开发了大量的生物信息学工具。笔者从测序技术、基因组装技术和全基因组测序生物信息学分析等方面,罗列了目前已完成全基因组测序的木本植物,介绍了全基因组测序技术的发展与应用,以及适用于第3代数据基因组组装的生物学分析软件,为林木基因组研究者提供一定的借鉴。     

Non-invasive prenatal measurement of the fetal genome

The vast majority of prenatal genetic testing requires invasive sampling. However, this poses a risk to the fetus, so one must make a decision that weighs the desire for genetic information against the risk of an adverse outcome due to hazards of the testing process. These issues are not required to be coupled, and it would be desirable to discover genetic information about the fetus without incurring a health risk. Here we demonstrate that it is possible to non-invasively sequence the entire prenatal genome. Our results show that molecular counting of parental haplotypes in maternal plasma by shotgun sequencing of maternal plasma DNA allows the inherited fetal genome to be deciphered non-invasively. We also applied the counting principle directly to each allele in the fetal exome by performing exome capture on maternal plasma DNA before shotgun sequencing. This approach enables non-invasive exome screening of clinically relevant and deleterious alleles that were paternally inherited or had arisen as de novo germline mutations, and complements the haplotype counting approach to provide a comprehensive view of the fetal genome. Non-invasive determination of the fetal genome may ultimately facilitate the diagnosis of all inherited and de novo genetic disease.     

Genomes for medicine

We have the human genome sequence. It is freely available, accurate and nearly complete. But is the genome ready for medicine? The new resource is already changing genetic research strategies to find information of medical value. Now we need high-quality annotation of all the functionally important sequences and the variations within them that contribute to health and disease. To achieve this, we need more genome sequences, systematic experimental analyses, and extensive information on human phenotypes. Flexible and user-friendly access to well-annotated genomes will create an environment for innovation, and the potential for unlimited use of sequencing in biomedical research and practice.     

A physical map of the mouse genome

A physical map of a genome is an essential guide for navigation, allowing the location of any gene or other landmark in the chromosomal DNA. We have constructed a physical map of the mouse genome that contains 296 contigs of overlapping bacterial clones and 16,992 unique markers. The mouse contigs were aligned to the human genome sequence on the basis of 51,486 homology matches, thus enabling use of the conserved synteny (correspondence between chromosome blocks) of the two genomes to accelerate construction of the mouse map. The map provides a framework for assembly of whole-genome shotgun sequence data, and a tile path of clones for generation of the reference sequence. Definition of the human-mouse alignment at this level of resolution enables identification of a mouse clone that corresponds to almost any position in the human genome. The human sequence may be used to facilitate construction of other mammalian genome maps using the same strategy.     

Mapping and sequencing of structural variation from eight human genomes

Genetic variation among individual humans occurs on many different scales, ranging from gross alterations in the human karyotype to single nucleotide changes. Here we explore variation on an intermediate scale--particularly insertions, deletions and inversions affecting from a few thousand to a few million base pairs. We employed a clone-based method to interrogate this intermediate structural variation in eight individuals of diverse geographic ancestry. Our analysis provides a comprehensive overview of the normal pattern of structural variation present in these genomes, refining the location of 1,695 structural variants. We find that 50% were seen in more than one individual and that nearly half lay outside regions of the genome previously described as structurally variant. We discover 525 new insertion sequences that are not present in the human reference genome and show that many of these are variable in copy number between individuals. Complete sequencing of 261 structural variants reveals considerable locus complexity and provides insights into the different mutational processes that have shaped the human genome. These data provide the first high-resolution sequence map of human structural variation--a standard for genotyping platforms and a prelude to future individual genome sequencing projects.     

黄连基因组勘测与分析

本研究基于下一代测序技术,对黄连基因组进行了勘测,构建了两个插入片段大小分别为200bp和500bp的文库,进行了深度约30X的测序。通过测序获得了54Gb的原始数据,过滤后得到44.8G数据。通过SOAP de nove软件组装后初步获得了contig和Scaffold序列,进一步分析结果显示其基因组大小为1,116Mb左右,大约具有1.1%的杂合度,说明要完成该物种的全基因测序可能在使用鸟枪法的同时,还应该联合BAC文库测序等多种方法.对这些数据进行了初步的组装,获得了130,381条scaffold序列.     

DNA fingerprinting in birds

Several regions of the human genome are highly variable in populations because the number of repeats in these regions of a short 'minisatellite' sequence varies at high frequency. Different minisatellites have a core sequence in common, however, and probes made up of tandem repeats of this core sequence detect many highly variable DNA fragments in several species including humans, cats, dogs and mice. The hypervariable sequences detected in this way are dispersed in the genome and their variability means that they can be used as a DNA 'fingerprint', providing a novel method for the identification of individuals, confirmation of biological relationships and human genetic analysis. We show here that human minisatellite-derived probes also detect highly variable regions in bird DNAs. Segregation analysis in a house sparrow family confirms that these regions comprise many mostly heterozygous dispersed loci and we conclude that house sparrow DNA fingerprints are analogous to those of humans. Fingerprint analysis identified one nestling, with fingerprint bands not present in the parent pair's fingerprints, which we conclude resulted from an extrapair copulation. Extrabond copulations have been described in many wild bird species, but their success and hence adaptive significance have rarely been quantifiable. DNA fingerprinting will be of great significance to studies of the sociobiology, demography and ecology of wild birds.     

A haplotype map of the human genome

Inherited genetic variation has a critical but as yet largely uncharacterized role in human disease. Here we report a public database of common variation in the human genome: more than one million single nucleotide polymorphisms (SNPs) for which accurate and complete genotypes have been obtained in 269 DNA samples from four populations, including ten 500-kilobase regions in which essentially all information about common DNA variation has been extracted. These data document the generality of recombination hotspots, a block-like structure of linkage disequilibrium and low haplotype diversity, leading to substantial correlations of SNPs with many of their neighbours. We show how the HapMap resource can guide the design and analysis of genetic association studies, shed light on structural variation and recombination, and identify loci that may have been subject to natural selection during human evolution.     

Accurate whole-genome sequencing and haplotyping from 10 to 20 human cells

Recent advances in whole-genome sequencing have brought the vision of personal genomics and genomic medicine closer to reality. However, current methods lack clinical accuracy and the ability to describe the context (haplotypes) in which genome variants co-occur in a cost-effective manner. Here we describe a low-cost DNA sequencing and haplotyping process, long fragment read (LFR) technology, which is similar to sequencing long single DNA molecules without cloning or separation of metaphase chromosomes. In this study, ten LFR libraries were made using only ～100?picograms of human DNA per sample. Up to 97% of the heterozygous single nucleotide variants were assembled into long haplotype contigs. Removal of false positive single nucleotide variants not phased by multiple LFR haplotypes resulted in a final genome error rate of 1 in 10?megabases. Cost-effective and accurate genome sequencing and haplotyping from 10-20 human cells, as demonstrated here, will enable comprehensive genetic studies and diverse clinical applications.     

Positive identification of an immigration test-case using human DNA fingerprints

The human genome contains a set of minisatellites, each of which consists of tandem repeats of a DNA segment containing the 'core' sequence, a putative recombination signal in human DNA. Multiallelic variation in the number of tandem repeats occurs at many of these minisatellite loci. Hybridization probes consisting of tandem repeats of the core sequence detect many hypervariable minisatellites simultaneously in human DNA, to produce a DNA fingerprint that is completely individual-specific and shows somatic and germline stability. These DNA fingerprints are derived from a large number of highly informative dispersed autosomal loci and are suitable for linkage analysis in man, and for individual identification in, for example, forensic science and paternity testing. They can also be used to resolve immigration disputes arising from lack of proof of family relationships. To illustrate the potential for positive or inclusive identification, we now describe the DNA fingerprint analysis of an immigration case, the resolution of which would have been very difficult and laborious using currently available single-locus genetic markers.     

Sequencing, annotation, and comparative analysis of nine BACs of the giant panda (Ailuropoda melanoleuca)

The BGI has carried out deep sequence exploration of nine BACs from the giant panda using traditional Sanger sequencing methods.