I am doing some analysis to identify differential accessible regions between two conditions using atac-seq. The workflow I have followed uses peak calling with macs2 and then differential analysis of these peaks with deseq2. This gave me good results with expected genes changing accessibility .
However, after reading more about atac-seq I am a little confused if my analysis is correct. As atac-seq produces both nfr and nucleosomal fragments, it seems that some workflows recommend filtering the bams for nfr fragments <100bp and then using them for further analysis. However, some workflows also use all reads without filtering which is what I did.
My question is if my current analysis makes sense or if I should repeat it after filtering my bams to <100bp fragments? I am also concerned that in that case I am discarding more than half of all my data. What is the best practice in this case? Also if I do the filtering, should I use the filtered bams for both peak calling and and deseq2 or for only one of these steps?