Replicates in Bisulfite data

Question

[chaudharyc61]

Hello everyone

I am working with plant BS-Seq data. I have six datasets, three mutant and three wild type. all the datasets are generated with different samples and experiments. So the replicates are of different experiments,same tissue.

My question is should i take the unique positions of Cytosine after merging or should i take common from all the replicates in each phenotype ? And if i take common sites there are some positions which might be important as the samples are different for making the replicates and removing those sites might not be helpful for the result. right ? Taking unique sites from all the replicates (per phenotype) means I am merging three replicates and taking all the positions. What to do ? I can't figure it out.

Please help me out...

Thanks Chandan Kumar

Answer 1

[dariober]

three mutant and three wild type.

I suspect you are going to look for differential methylation. If so, have a look at this method based on RNAseq analysis Differential methylation analysis of reduced representation bisulfite sequencing experiments using edgeR

Regarding which cytosines to consider, I would keep positions where at least 5 of the samples have decent coverage (where "decent" depends on the data you have...).