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| Status |
Public on Jun 25, 2011 |
| Title |
Application of Microdroplet PCR for Large-Scale Targeted Bisulfite Sequencing |
| Organism |
Homo sapiens |
| Experiment type |
Methylation profiling by high throughput sequencing
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| Summary |
Cytosine methylation of DNA CpG dinucleotides in gene promoters is an epigenetic modification that regulates gene transcription. While many methods exist to interrogate methylation states, no current methods offer large-scale, targeted, single CpG resolution. We report an approach combining bisulfite treatment followed by RainDance microdroplet PCR with next-generation sequencing to assay the methylation state of 50 genes in the regions 1 kb upstream and downstream of their transcription start sites.
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| Overall design |
Wildtype and hypermethylated Jurkat DNA (New Englad Biolabs) was treated with bisulfite to convert all unmethylated cytosines to uracil. Following bisulfite treatment, targeted amplification was carried out using a custom primer library and microdroplet PCR. PCR product was sheared to 200 bp and ligated to sequencing adapters following standard protocols. Sequencing was conducted with single-end 100 bp reads on an Illumina GAIIx for wild type Jurkat DNA or Jurkat CpG DNA with a single sample per lane.
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| Contributor(s) |
Komori HK, LaMere SA, Torkamani A, Hart GT, Kotsopoulos S, Warner J, Samuels ML, Olson J, Head SR, Ordoukhanian P, Lee PL, Link DR, Salomon DR |
| Citation(s) |
21757609 |
| Submission date |
Jun 23, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Traver Hart |
| E-mail(s) |
traver.hart@gmail.com
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| Organization name |
The Scripps Research Institute
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| Department |
Molecular & Experimental Medicine
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| Lab |
Salomon Lab
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| Street address |
10550 North Torrey Pines Road, MEM241B
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| City |
La Jo |
| State/province |
CA |
| ZIP/Postal code |
92037 |
| Country |
USA |
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| Platforms (1) |
| GPL9115 |
Illumina Genome Analyzer II (Homo sapiens) |
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| Samples (2) |
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| Relations |
| SRA |
SRP007358 |
| BioProject |
PRJNA143939 |
