Abstract:
:The relatively short read lengths associated with the most cost-effective DNA sequencing technologies have limited their use in de novo genome assembly, structural variation detection, and haplotype-resolved genome sequencing. Consequently, there is a strong need for methods that capture various scales of contiguity information at a throughput commensurate with the current scale of massively parallel sequencing. We propose in situ library construction and optical sequencing on the flow cells of currently available massively parallel sequencing platforms as an efficient means of capturing both contiguity information and primary sequence with a single technology. In this proof-of-concept study, we demonstrate basic feasibility by generating >30,000 Escherichia coli paired-end reads separated by 1, 2, or 3 kb using in situ library construction on standard Illumina flow cells. We also show that it is possible to stretch single molecules ranging from 3 to 8 kb on the surface of a flow cell before in situ library construction, thereby enabling the production of clusters whose physical relationship to one another on the flow cell is related to genomic distance.
journal_name
Proc Natl Acad Sci U S Aauthors
Schwartz JJ,Lee C,Hiatt JB,Adey A,Shendure Jdoi
10.1073/pnas.1202680109subject
Has Abstractpub_date
2012-11-13 00:00:00pages
18749-54issue
46eissn
0027-8424issn
1091-6490pii
1202680109journal_volume
109pub_type
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