Accurate whole human genome sequencing using reversible terminator chemistry.

DNA sequence information underpins genetic research, enabling discoveries of important biological or medical benefit. Sequencing projects have traditionally used long (400-800 base pair) 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 intraspecies genetic variation. Here 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 characterize four million single-nucleotide polymorphisms and four hundred thousand structural variants, many of which were previously unknown. Our approach is effective for accurate, rapid and economical whole-genome re-sequencing and many other biomedical applications.

Whole-genome re-sequencing.

DNA sequencing can be used to gain important information on genes, genetic variation and gene function for biological and medical studies. The growing collection of publicly available reference genome sequences will underpin a new era of whole genome re-sequencing, but sequencing costs need to fall and throughput needs to rise by several orders of magnitude. Novel technologies are being developed to meet this need by generating massive amounts of sequence that can be aligned to the reference sequence. The challenge is to maintain the high standards of accuracy and completeness that are hallmarks of the previous genome projects. One or more new sequencing technologies are expected to become the mainstay of future research, and to make DNA sequencing centre stage as a routine tool in genetic research in the coming years.

The fine-scale structure of recombination rate variation in the human genome.

The nature and scale of recombination rate variation are largely unknown for most species. In humans, pedigree analysis has documented variation at the chromosomal level, and sperm studies have identified specific hotspots in which crossing-over events cluster. To address whether this picture is representative of the genome as a whole, we have developed and validated a method for estimating recombination rates from patterns of genetic variation. From extensive single-nucleotide polymorphism surveys in European and African populations, we find evidence for extreme local rate variation spanning four orders in magnitude, in which 50% of all recombination events take place in less than 10% of the sequence. We demonstrate that recombination hotspots are a ubiquitous feature of the human genome, occurring on average every 200 kilobases or less, but recombination occurs preferentially outside genes.