Targeted bisulfite sequencing by solution hybrid selection and massively parallel sequencing

Hdl Handle:
http://hdl.handle.net/10675.2/761
Title:
Targeted bisulfite sequencing by solution hybrid selection and massively parallel sequencing
Authors:
Lee, Eun-Joon; Pei, Lirong; Srivastava, Gyan; Joshi, Trupti; Kushwaha, Garima; Choi, Jeong-Hyeon; Robertson, Keith D.; Wang, Xinguo; Colbourne, John K. ( 0000-0002-6966-2972 ) ; Zhang, Lu; Schroth, Gary P.; Xu, Dong; Zhang, Kun; Shi, Huidong
Abstract:
We applied a solution hybrid selection approach to the enrichment of CpG islands (CGIs) and promoter sequences from the human genome for targeted high-throughput bisulfite sequencing. A single lane of Illumina sequences allowed accurate and quantitative analysis of ~1 million CpGs in more than 21â 408 CGIs and more than 15â 946 transcriptional regulatory regions. Of the CpGs analyzed, 77â 84% fell on or near capture probe sequences; 69â 75% fell within CGIs. More than 85% of capture probes successfully yielded quantitative DNA methylation information of targeted regions. Differentially methylated regions (DMRs) were identified in the 5â ²-end regulatory regions, as well as the intra- and intergenic regions, particularly in the X-chromosome among the three breast cancer cell lines analyzed. We chose 46 candidate loci (762 CpGs) for confirmation with PCR-based bisulfite sequencing and demonstrated excellent correlation between two data sets. Targeted bisulfite sequencing of three DNA methyltransferase (DNMT) knockout cell lines and the wild-type HCT116 colon cancer cell line revealed a significant decrease in CpG methylation for the DNMT1 knockout and DNMT1, 3B double knockout cell lines, but not in DNMT3B knockout cell line. We demonstrated the targeted bisulfite sequencing approach to be a powerful method to uncover novel aberrant methylation in the cancer epigenome. Since all targets were captured and sequenced as a pool through a series of single-tube reactions, this method can be easily scaled up to deal with a large number of samples.
Citation:
Nucleic Acids Res. 2011 Oct 23; 39(19):e127
Issue Date:
23-Oct-2011
URI:
http://hdl.handle.net/10675.2/761
DOI:
10.1093/nar/gkr598
PubMed ID:
21785137
PubMed Central ID:
PMC3201883
Type:
Article
ISSN:
1362-4962
Appears in Collections:
Georgia Cancer Center: Faculty Research and Presentations

Full metadata record

DC FieldValue Language
dc.contributor.authorLee, Eun-Joonen_US
dc.contributor.authorPei, Lirongen_US
dc.contributor.authorSrivastava, Gyanen_US
dc.contributor.authorJoshi, Truptien_US
dc.contributor.authorKushwaha, Garimaen_US
dc.contributor.authorChoi, Jeong-Hyeonen_US
dc.contributor.authorRobertson, Keith D.en_US
dc.contributor.authorWang, Xinguoen_US
dc.contributor.authorColbourne, John K.en_US
dc.contributor.authorZhang, Luen_US
dc.contributor.authorSchroth, Gary P.en_US
dc.contributor.authorXu, Dongen_US
dc.contributor.authorZhang, Kunen_US
dc.contributor.authorShi, Huidongen_US
dc.date.accessioned2012-10-26T20:30:40Z-
dc.date.available2012-10-26T20:30:40Z-
dc.date.issued2011-10-23en_US
dc.identifier.citationNucleic Acids Res. 2011 Oct 23; 39(19):e127en_US
dc.identifier.issn1362-4962en_US
dc.identifier.pmid21785137en_US
dc.identifier.doi10.1093/nar/gkr598en_US
dc.identifier.urihttp://hdl.handle.net/10675.2/761-
dc.description.abstractWe applied a solution hybrid selection approach to the enrichment of CpG islands (CGIs) and promoter sequences from the human genome for targeted high-throughput bisulfite sequencing. A single lane of Illumina sequences allowed accurate and quantitative analysis of ~1 million CpGs in more than 21â 408 CGIs and more than 15â 946 transcriptional regulatory regions. Of the CpGs analyzed, 77â 84% fell on or near capture probe sequences; 69â 75% fell within CGIs. More than 85% of capture probes successfully yielded quantitative DNA methylation information of targeted regions. Differentially methylated regions (DMRs) were identified in the 5â ²-end regulatory regions, as well as the intra- and intergenic regions, particularly in the X-chromosome among the three breast cancer cell lines analyzed. We chose 46 candidate loci (762 CpGs) for confirmation with PCR-based bisulfite sequencing and demonstrated excellent correlation between two data sets. Targeted bisulfite sequencing of three DNA methyltransferase (DNMT) knockout cell lines and the wild-type HCT116 colon cancer cell line revealed a significant decrease in CpG methylation for the DNMT1 knockout and DNMT1, 3B double knockout cell lines, but not in DNMT3B knockout cell line. We demonstrated the targeted bisulfite sequencing approach to be a powerful method to uncover novel aberrant methylation in the cancer epigenome. Since all targets were captured and sequenced as a pool through a series of single-tube reactions, this method can be easily scaled up to deal with a large number of samples.en_US
dc.rights© The Author(s) 2011. Published by Oxford University Press.en_US
dc.subjectMethods Onlineen_US
dc.titleTargeted bisulfite sequencing by solution hybrid selection and massively parallel sequencingen_US
dc.typeArticleen_US
dc.identifier.pmcidPMC3201883en_US
dc.contributor.corporatenameGHSU Cancer Center-
dc.contributor.corporatenameDepartment of Biochemistry and Molecular Biology-
dc.contributor.corporatenameDepartment of Biostatistics and Epidemiology-

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