Capturing native long-range contiguity by in situ library construction and optical sequencing.
Proc Natl Acad Sci U S A
; 109(46): 18749-54, 2012 Nov 13.
Article
em En
| MEDLINE
| ID: mdl-23112150
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.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA Bacteriano
/
Biblioteca Genômica
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Genoma Bacteriano
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Análise de Sequência de DNA
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Escherichia coli
Idioma:
En
Revista:
Proc Natl Acad Sci U S A
Ano de publicação:
2012
Tipo de documento:
Article
País de afiliação:
Estados Unidos