Format

Send to

Choose Destination
DNA Res. 2015 Feb;22(1):13-8. doi: 10.1093/dnares/dsu034. Epub 2014 Oct 16.

Highly sensitive targeted methylome sequencing by post-bisulfite adaptor tagging.

Author information

1
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113-0033, Japan Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Higashi-ku, Fukuoka 812-8582, Japan Department of Biochemistry, Kyushu University Graduate School of Medical Sciences, Higashi-ku, Fukuoka 812-8582, Japan.
2
Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Bunkyo-ku, Tokyo 113-0033, Japan Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency (JST), Higashi-ku, Fukuoka 812-8582, Japan Department of Biochemistry, Kyushu University Graduate School of Medical Sciences, Higashi-ku, Fukuoka 812-8582, Japan tito@med.kyushu-u.ac.jp.

Abstract

The current gold standard method for methylome analysis is whole-genome bisulfite sequencing (WGBS), but its cost is substantial, especially for the purpose of multi-sample comparison of large methylomes. Shotgun bisulfite sequencing of target-enriched DNA, or targeted methylome sequencing (TMS), can be a flexible, cost-effective alternative to WGBS. However, the current TMS protocol requires a considerable amount of input DNA and hence is hardly applicable to samples of limited quantity. Here we report a method to overcome this limitation by using post-bisulfite adaptor tagging (PBAT), in which adaptor tagging is conducted after bisulfite treatment to circumvent bisulfite-induced loss of intact sequencing templates, thereby enabling TMS of a 100-fold smaller amount of input DNA with far fewer cycles of polymerase chain reaction than in the current protocol. We thus expect that the PBAT-mediated TMS will serve as an invaluable method in epigenomics.

KEYWORDS:

DNA methylation; massively parallel sequencing; target enrichment

PMID:
25324297
PMCID:
PMC4379973
DOI:
10.1093/dnares/dsu034
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Silverchair Information Systems Icon for PubMed Central
Loading ...
Support Center