DNA sequence information underpins genetic research, enabling discoveries of important biological or medical benefit. Sequencing projects have traditionally employed long (400–800 bp) reads, but the existence of reference sequences for the human and many other genomes makes it possible to develop new, fast approaches to re-sequencing, whereby shorter reads are compared to a reference to identify intra-species genetic variation. We report an approach that generates several billion bases of accurate nucleotide sequence per experiment at low cost. Single molecules of DNA are attached to a flat surface, amplified
in situ
and used as templates for synthetic sequencing with fluorescent reversible terminator deoxyribonucleotides. Images of the surface are analysed to generate high quality sequence. We demonstrate application of this approach to human genome sequencing on flow-sorted X chromosomes and then scale the approach to determine the genome sequence of a male Yoruba from Ibadan, Nigeria. We build an accurate consensus sequence from >30x average depth of paired 35-base reads. We characterise four million SNPs and four hundred thousand structural variants, many of which are previously unknown. Our approach is effective for accurate, rapid and economical whole genome re-sequencing and many other biomedical applications.
Siberian taimen Hucho taimen is the largest representative of the family Salmonidae inhabiting rivers of northern Eurasia. The species is under intensive aquaculture activity. To monitor natural taimen populations we have sequenced a portion (8,141 bp) of the mitochondrial (mt) genome in 28 specimens of H. taimen from six localities in the Amur River basin. Nucleotide variability is low (π = 0.0010), but structured in two divergent haplotype groups. A comparison of the data with the GenBank H. taimen mt genome (HQ897271) reveals significant differences between them in spite of the fact that the fish specimens come from neighboring geographical areas. The distribution of divergence is non-uniform with two highly pronounced divergent regions centered on two genes, ND3 and ND6. To clarify the pattern of divergence we sequenced the corresponding portion of the mt genome of lenok Brachymystax tumensis and analyzed the GenBank complete mt genomes of related species. We have found that the first and second divergent regions are identical between the GenBank H. taimen and two lenok subspecies, B. lenok and B. lenok tsinlingensis, respectively. Consequently, both divergent regions represent introgressed mtDNA resulting from intergeneric hybridization between the two lenok subspecies and H. taimen. Introgression is, however, not detected in our specimens. This plus the precise identity of the introgressed fragments between the donor and the recipient GenBank sequence suggests that the introgression is local and very recent, probably due to artificial manipulations involving taimen – lenok intergeneric hybridization. Human-mediated hybridization may become a major threat to aquatic biodiversity. Consequently we suggest that due attention needs to be given to this threat by means of responsible breeding program management, so as to prevent a potential spread of hybrid fishes that could jeopardize the resilience of locally adapted gene pools of the native H. taimen populations.
Chars of the genus Salvelinus , inhabiting lakes and lake-river systems, comprise morphological and ecological forms whose taxonomic status is under dispute. In the present work, we have examined genetic variation and divergence in various chars from the Kronotsky lake basin: the lacustrine chars (white, nose, and longhead) and riverine Dolly Varden Salvelinus malma. The study was conducted using analysis of allozyme and microsatellite loci, myogens, RAPD, and restriction analysis of two mtDNA segments. The estimates of heterozygosity at allozyme and microsatellite loci were similar to the corresponding parameters in populations of northern Dolly Varden and Arctic char. Heterozygote deficit was recorded in both samples of separate forms, and in the total sample of all chars from Kronotsky Lake. For allozyme and microsatellite loci, appreciable genetic differentiation among the samples of different char forms was found, which was comparable to that among the spatially isolated populations of northern Dolly Varden. This result indicates reproductive isolation among the char forms examined. However, this isolation is not complete, because no fixed differences between the forms by any of the genetic systems analyzed were found. The genetic differentiation among different forms of lacustrine chars, which corresponds to the interpopulation rather than interspecific level, is thought to be explained by their comparatively recent divergence.
PLANT GENETICS
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