Bismark reads failing to align

Question

[Anna S]

Hello,

I'm using Bismark for the first time, and 99.99% of the reads are failing to align for my paired dataset of seq length 51 bp. I have tried using the default -l, as well as -l of 40, the default -n as well as -n of 1, the original fastq files as well as quality trimmed and adaptor cut ones, etc., but none of these variations have made much of a difference. The FastQC output shows that the per base and per sequence quality scores are very good. Do you have any suggestions on how to proceed?

Thank you!!

Anna

[Anna S]

Please delete this question as it turns out to have been my misunderstanding of the datasets (2 lanes of the same end files vs paired-end files). Bismark is working great now!!

Answer 1

[dariober]

Without more detail is hard to tell where the problem is... Assuming the reference genome is the correct one, I would try to align only read 1 and see if the mapping goes up. Also make sure the paired reads have been trimmed in such way that the two fastq files in a pair remain in sync (i.e. by using trim_galore)