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2017 ; 27
(5
): 757-767
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Direct determination of diploid genome sequences
#MMPMID28381613
Weisenfeld NI
; Kumar V
; Shah P
; Church DM
; Jaffe DB
Genome Res
2017[May]; 27
(5
): 757-767
PMID28381613
show ga
Determining the genome sequence of an organism is challenging, yet fundamental to
understanding its biology. Over the past decade, thousands of human genomes have
been sequenced, contributing deeply to biomedical research. In the vast majority
of cases, these have been analyzed by aligning sequence reads to a single
reference genome, biasing the resulting analyses, and in general, failing to
capture sequences novel to a given genome. Some de novo assemblies have been
constructed free of reference bias, but nearly all were constructed by merging
homologous loci into single "consensus" sequences, generally absent from nature.
These assemblies do not correctly represent the diploid biology of an individual.
In exactly two cases, true diploid de novo assemblies have been made, at great
expense. One was generated using Sanger sequencing, and one using thousands of
clone pools. Here, we demonstrate a straightforward and low-cost method for
creating true diploid de novo assemblies. We make a single library from ?1 ng of
high molecular weight DNA, using the 10x Genomics microfluidic platform to
partition the genome. We applied this technique to seven human samples,
generating low-cost HiSeq X data, then assembled these using a new "pushbutton"
algorithm, Supernova. Each computation took 2 d on a single server. Each yielded
contigs longer than 100 kb, phase blocks longer than 2.5 Mb, and scaffolds longer
than 15 Mb. Our method provides a scalable capability for determining the actual
diploid genome sequence in a sample, opening the door to new approaches in
genomic biology and medicine.
free
free
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