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.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 18749-18754 |
| Number of pages | 6 |
| Journal | Proceedings of the National Academy of Sciences of the United States of America |
| Volume | 109 |
| Issue number | 46 |
| DOIs | |
| State | Published - Nov 13 2012 |
| Externally published | Yes |
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Keywords
- Jumping reads
- Molecular biophysics
- Transposase
ASJC Scopus subject areas
- General
Cite this
Capturing native long-range contiguity by in situ library construction and optical sequencing. / Schwartz, Jerrod J.; Lee, Choli; Hiatt, Joseph B.; Adey, Andrew; Shendure, Jay.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 109, No. 46, 13.11.2012, p. 18749-18754.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Capturing native long-range contiguity by in situ library construction and optical sequencing
AU - Schwartz, Jerrod J.
AU - Lee, Choli
AU - Hiatt, Joseph B.
AU - Adey, Andrew
AU - Shendure, Jay
PY - 2012/11/13
Y1 - 2012/11/13
N2 - 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.
AB - 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.
KW - Jumping reads
KW - Molecular biophysics
KW - Transposase
UR - http://www.scopus.com/inward/record.url?scp=84869234142&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84869234142&partnerID=8YFLogxK
U2 - 10.1073/pnas.1202680109
DO - 10.1073/pnas.1202680109
M3 - Article
C2 - 23112150
AN - SCOPUS:84869234142
VL - 109
SP - 18749
EP - 18754
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
SN - 0027-8424
IS - 46
ER -