Initial sequencing and analysis of the human genome.
TL;DR: The results of an international collaboration to produce and make freely available a draft sequence of the human genome are reported and an initial analysis is presented, describing some of the insights that can be gleaned from the sequence.
Abstract: The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.
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TL;DR: The definition and use of family-specific, manually curated gathering thresholds are explained and some of the features of domains of unknown function (also known as DUFs) are discussed, which constitute a rapidly growing class of families within Pfam.
Abstract: Pfam is a widely used database of protein families and domains. This article describes a set of major updates that we have implemented in the latest release (version 24.0). The most important change is that we now use HMMER3, the latest version of the popular profile hidden Markov model package. This software is approximately 100 times faster than HMMER2 and is more sensitive due to the routine use of the forward algorithm. The move to HMMER3 has necessitated numerous changes to Pfam that are described in detail. Pfam release 24.0 contains 11,912 families, of which a large number have been significantly updated during the past two years. Pfam is available via servers in the UK (http://pfam.sanger.ac.uk/), the USA (http://pfam.janelia.org/) and Sweden (http://pfam.sbc.su.se/).
14,075 citations
TL;DR: Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems are indicated.
Abstract: A 2.91-billion base pair (bp) consensus sequence of the euchromatic portion of the human genome was generated by the whole-genome shotgun sequencing method. The 14.8-billion bp DNA sequence was generated over 9 months from 27,271,853 high-quality sequence reads (5.11-fold coverage of the genome) from both ends of plasmid clones made from the DNA of five individuals. Two assembly strategies-a whole-genome assembly and a regional chromosome assembly-were used, each combining sequence data from Celera and the publicly funded genome effort. The public data were shredded into 550-bp segments to create a 2.9-fold coverage of those genome regions that had been sequenced, without including biases inherent in the cloning and assembly procedure used by the publicly funded group. This brought the effective coverage in the assemblies to eightfold, reducing the number and size of gaps in the final assembly over what would be obtained with 5.11-fold coverage. The two assembly strategies yielded very similar results that largely agree with independent mapping data. The assemblies effectively cover the euchromatic regions of the human chromosomes. More than 90% of the genome is in scaffold assemblies of 100,000 bp or more, and 25% of the genome is in scaffolds of 10 million bp or larger. Analysis of the genome sequence revealed 26,588 protein-encoding transcripts for which there was strong corroborating evidence and an additional approximately 12,000 computationally derived genes with mouse matches or other weak supporting evidence. Although gene-dense clusters are obvious, almost half the genes are dispersed in low G+C sequence separated by large tracts of apparently noncoding sequence. Only 1.1% of the genome is spanned by exons, whereas 24% is in introns, with 75% of the genome being intergenic DNA. Duplications of segmental blocks, ranging in size up to chromosomal lengths, are abundant throughout the genome and reveal a complex evolutionary history. Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems. DNA sequence comparisons between the consensus sequence and publicly funded genome data provided locations of 2.1 million single-nucleotide polymorphisms (SNPs). A random pair of human haploid genomes differed at a rate of 1 bp per 1250 on average, but there was marked heterogeneity in the level of polymorphism across the genome. Less than 1% of all SNPs resulted in variation in proteins, but the task of determining which SNPs have functional consequences remains an open challenge.
12,098 citations
TL;DR: In a four-genome analysis of 3' UTRs, approximately 13,000 regulatory relationships were detected above the estimate of false-positive predictions, thereby implicating as miRNA targets more than 5300 human genes, which represented 30% of the gene set.
11,624 citations
TL;DR: A mature web tool for rapid and reliable display of any requested portion of the genome at any scale, together with several dozen aligned annotation tracks, is provided at http://genome.ucsc.edu.
Abstract: As vertebrate genome sequences near completion and research refocuses to their analysis, the issue of effective genome annotation display becomes critical. A mature web tool for rapid and reliable display of any requested portion of the genome at any scale, together with several dozen aligned annotation tracks, is provided at http://genome.ucsc.edu. This browser displays assembly contigs and gaps, mRNA and expressed sequence tag alignments, multiple gene predictions, cross-species homologies, single nucleotide polymorphisms, sequence-tagged sites, radiation hybrid data, transposon repeats, and more as a stack of coregistered tracks. Text and sequence-based searches provide quick and precise access to any region of specific interest. Secondary links from individual features lead to sequence details and supplementary off-site databases. One-half of the annotation tracks are computed at the University of California, Santa Cruz from publicly available sequence data; collaborators worldwide provide the rest. Users can stably add their own custom tracks to the browser for educational or research purposes. The conceptual and technical framework of the browser, its underlying MYSQL database, and overall use are described. The web site currently serves over 50,000 pages per day to over 3000 different users.
9,605 citations
TL;DR: Velvet represents a new approach to assembly that can leverage very short reads in combination with read pairs to produce useful assemblies and is in close agreement with simulated results without read-pair information.
Abstract: We have developed a new set of algorithms, collectively called "Velvet," to manipulate de Bruijn graphs for genomic sequence assembly. A de Bruijn graph is a compact representation based on short words (k-mers) that is ideal for high coverage, very short read (25-50 bp) data sets. Applying Velvet to very short reads and paired-ends information only, one can produce contigs of significant length, up to 50-kb N50 length in simulations of prokaryotic data and 3-kb N50 on simulated mammalian BACs. When applied to real Solexa data sets without read pairs, Velvet generated contigs of approximately 8 kb in a prokaryote and 2 kb in a mammalian BAC, in close agreement with our simulated results without read-pair information. Velvet represents a new approach to assembly that can leverage very short reads in combination with read pairs to produce useful assemblies.
9,389 citations
References
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TL;DR: An organization of human 5 S rDNA repeats is inferred from Southern analyses of restriction digests of genomic DNA fractionated by pulsed-field and conventional gel electrophoreses and four clones have been partially characterized and shown to be 5 S sequences from loci separate from the tandem repeat units.
89 citations
TL;DR: The genome of the fruitfly Drosophila melanogaster is 180 megabases, whereas that of the European brown grasshopper Podisma pedestris is 18,000 Mb, a difference in genome size of a factor of 100.
Abstract: For 50 years now, one of the enigmas of molecular evolution has been the C-value paradox, which refers to the often massive, counterintuitive and seemingly arbitrary differences in genome size observed among eukaryotic organisms. For example, the genome of the fruitfly Drosophila melanogaster is 180 megabases (Mb), whereas that of the European brown grasshopper Podisma pedestris is 18,000 Mb. The difference in genome size of a factor of 100 is difficult to explain in view of the apparently similar levels of evolutionary, developmental and behavioural complexity of these organisms.
88 citations
TL;DR: It is proposed that the distal CMT1A-REP represents the progenitor copy of COX10 exon VI which was duplicated with surrounding intronic sequences during mammalian genome evolution and that the HNPP deletion results in aCOX10 null allele.
Abstract: The CMT1A-REPs are two large directly repeating DNA sequences located on chromosome 17p11.2-p12 flanking the region duplicated in patients with Charcot-Marie-Tooth disease type 1A (CMT1A) and deleted in patients with hereditary neuropathy with liability to pressure palsies (HNPP). We have sequenced two cosmids, c74F4 and c15H12, which contain the entire proximal and distal CMT1A-REPs and determined that these repeats are approximately 99% identical across a 24,011 bp region. In addition, both contain an exon of the human heme A:farnesyltransferase gene (COX10). Hybridization studies revealed that COX10 spans the distal CMT1A-REP, while the proximal CMT1A-REP contains an isolated COX10 'pseudo-exon'. There is also a COX10 hybridization signal on chromosome 10 which appears to represent a processed pseudogene. We propose that the distal CMT1A-REP represents the progenitor copy of COX10 exon VI which was duplicated with surrounding intronic sequences during mammalian genome evolution and that the HNPP deletion results in a COX10 null allele.
88 citations
TL;DR: The analysis of spacer DNA by molecular cloning and its organization in the genome by Southern transfer analysis reveal both length and sequence variation of the spacer.
Abstract: The complete repeating unit of the human ribosomal RNA gene has been reconstructed by the cloning of approximately 27 kilobases (kb) of non-transcribed spacer. The structure of this tandemly repeated gene can now be studied in its entirety. We report the analysis of spacer DNA by molecular cloning and its organization in the genome by Southern transfer analysis. These studies reveal both length and sequence variation of the spacer. Sequence variations are distributed throughout the spacer while the length variations exist near the 5' end of the transcript and just beyond the 3' end. The human spacer shares extensive homology with primates but little with other mammals. Within the primates the degree of homology reflects the rapid evolutionary changes characteristic of the primate group.
87 citations
TL;DR: It is proposed that AASS catalyzes the first two steps of the major lysine-degradation pathway in human cells and that inactivating mutations in the AASS gene are a cause of hyperlysinemia.
Abstract: The first two steps in the mammalian lysine-degradation pathway are catalyzed by lysine-ketoglutarate reductase and saccharopine dehydrogenase, respectively, resulting in the conversion of lysine to α-aminoadipic semialdehyde Defects in one or both of these activities result in familial hyperlysinemia, an autosomal recessive condition characterized by hyperlysinemia, lysinuria, and variable saccharopinuria In yeast, lysine-ketoglutarate reductase and saccharopine dehydrogenase are encoded by the LYS1 and LYS9 genes, respectively, and we searched the available sequence databases for their human homologues We identified a single cDNA that encoded an apparently bifunctional protein, with the N-terminal half similar to that of yeast LYS1 and with the C-terminal half similar to that of yeast LYS9 This bifunctional protein has previously been referred to as "α-aminoadipic semialdehyde synthase," and we have tentatively designated this gene " AASS " The AASS cDNA contains an open reading frame of 2,781 bp predicted to encode a 927-amino-acid-long protein The gene has been sequenced and contains 24 exons scattered over 68 kb and maps to chromosome 7q313 Northern blot analysis revealed the presence of several transcripts in all tissues examined, with the highest expression occurring in the liver We sequenced the genomic DNA from a single patient with hyperlysinemia (JJa) The patient is the product of a consanguineous mating and is homozygous for an out-of-frame 9-bp deletion in exon 15, which results in a premature stop codon at position 534 of the protein On the basis of these and other results, we propose that AASS catalyzes the first two steps of the major lysine-degradation pathway in human cells and that inactivating mutations in the AASS gene are a cause of hyperlysinemia
87 citations