I have some RNA-seq data, in which some samples were paired-end reads, and some were single-end reads. I used Combat-seq from sva package to remove the batch effect with input is raw counts. Then I used the adjusted counts (after Combat-seq) as input for DESeq2 and made PCA plot (rld).
I also compiled some other RNA-seq data (paired-end) from 3 papers (I make a raw-count file again from their raw RNA-seq). Now, I set 5 batches: 1: my paired-end data, 2 my single-end data, 3-5: data from 3 different papers. Is that fine?
I am trying to use other methods of batch effect removal and compare those. I tried using the limma::removeBatchEffect. I did make dds file from DESeq2 (input is raw counts) => vsd (<-vst function) => limma::removeBatchEffect => PCA plot.
I had 2 PCA plot show totally differently by 2 different methods of batch effect removal.
I am so new to bioinformatics. Please give me some advice or share with me some relevant materials.
Thank you in advance!
