Want the sensitivity of WGSBS without the cost? RRBS brings down the scale and cost of WGSBS by only sequencing a reduced, representative sample of the whole genome. First the DNA is digested, typically with MspI which is methylation insensitive and cuts at CCGG sites. This enriches for CpG rich regions of the genome. The fragment ends are then filled in and adapters are ligated. Next the fragments are size selected, bisulfite converted, and sequenced.
This simple approach captures 85% of CpG islands and 60% of promoters and requires very little input sample (Gu et al., 2011). The reduction in DNA also means that fewer reads are needed for accurate sequencing, reducing cost dramatically. The obvious disadvantage of RRBS is that it doesn’t capture all CpG islands or promoters. Depending on the experiment, this can be major problem. Introduced in 2005 (Meissner et al., 2005) RRBS has become popular when high-throughput, low cost methylation analysis is needed such as clinical applications (Lee et al., 2014).
RRBS Additional Reading
This review covers many methylation analysis techniques including RRBS. The authors focus on the pros and cons of RRBS, technical challenges, and interesting insights for studies that have used it.
For more info about reduced representation bisulfite sequencing, check out this nice article on using RRBS for DNA methylation analysis from our friends at Active Motif.
- Gu, H., Smith, Z.D., Bock, C., Boyle, P., Gnirke, A., and Meissner, A. (2011). Preparation of reduced representation bisulfite sequencing libraries for genome-scale DNA methylation profiling. Nat. Protoc. 6, 468-481.
- Lee, Y.K., Jin, S., Duan, S., Lim, Y.C., Ng, D.P., Lin, X.M., Yeo, G.S., and Ding, C. (2014). Improved reduced representation bisulfite sequencing for epigenomic profiling of clinical samples. Biol. Proced. Online 16, 1-9222-16-1.
- Meissner, A., Gnirke, A., Bell, G.W., Ramsahoye, B., Lander, E.S., and Jaenisch, R. (2005). Reduced representation bisulfite sequencing for comparative high-resolution DNA methylation analysis. Nucleic Acids Res. 33, 5868-5877.