Hi all,
Looking for some advice. I have a small number of iPSC-derived oligodendrocytes (approx 10,000 cells/sample) that I'd like to do RNAseq and whole genome wide methylation analysis (e.g. Illumina 850k). Can anyone give me any insight as to whether this is feasible?
Does anyone have any experience of doing such work on small number of cells (and perhaps can recommend kits for extraction) and/or amplification steps if required?
Many thanks for any advice,
Vicky
Hi,
I have some experience of EWAS and RNA-Seq before but I seldom do expriment. I would do both RNA-Seq & EWAS on the same sample if I wanted to explain the difference of RNA level by DNA methylation. Make sure you have enough sample size to do EWAS first(34 paired in my case but less is ok). However, that's gonna cost too much since sometime only a small number of genes overlap in both EWAS and RNA-Seq. My point is, if you want to focus on RNA level, you can select the top interested gene by RNA-Seq on small sample size and the just use bisulfate sequencing to detect the DNA methylation of those gene; If you are interested in epigenetic you might do EWAS first and select the top CpG site to detect the mRNA level by PCR.
BTW, if you want to use 850k, make sure your DNA concentration > 3ug (in my case, 60－1000 ng/μL, OD260/280 1.7～2.1) and minimize the batch effect.
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