A MBD-seq protocol for large-scale methylome-wide studies with (very) low amounts of DNA.
|Title||A MBD-seq protocol for large-scale methylome-wide studies with (very) low amounts of DNA.|
|Publication Type||Journal Article|
|Year of Publication||2017|
|Authors||Aberg KA, Chan RF, Shabalin AA, Zhao M, Turecki G, Staunstrup NHeine, Starnawska A, Mors O, Xie LY, van den Oord EJcg|
|Date Published||2017 Jul 13|
We recently showed that, after optimization, our methyl-CpG binding domain sequencing (MBD-seq) application approximates the methylome-wide coverage obtained with whole-genome bisulfite sequencing (WGB-seq), but at a cost that enables adequately powered large-scale association studies. A prior drawback of MBD-seq is the relatively large amount of genomic DNA (ideally >1 µg) required to obtain high-quality data. Biomaterials are typically expensive to collect, provide a finite amount of DNA, and may simply not yield sufficient starting material. The ability to use low amounts of DNA will increase the breadth and number of studies that can be conducted. Therefore, we further optimized the enrichment step. With this low starting material protocol, MBD-seq performed equally well, or better, than the protocol requiring ample starting material (>1 µg). Using only 15 ng of DNA as input, there is minimal loss in data quality, achieving 93% of the coverage of WGB-seq (with standard amounts of input DNA) at similar false/positive rates. Furthermore, across a large number of genomic features, the MBD-seq methylation profiles closely tracked those observed for WGB-seq with even slightly larger effect sizes. This suggests that MBD-seq provides similar information about the methylome and classifies methylation status somewhat more accurately. Performance decreases with <15 ng DNA as starting material but, even with as little as 5 ng, MBD-seq still achieves 90% of the coverage of WGB-seq with comparable genome-wide methylation profiles. Thus, the proposed protocol is an attractive option for adequately powered and cost-effective methylome-wide investigations using (very) low amounts of DNA.
|Grant List||R01 MH099110 / MH / NIMH NIH HHS / United States |
R01 MH104576 / MH / NIMH NIH HHS / United States
R01 MH109525 / MH / NIMH NIH HHS / United States