Does HISAT-3N work for paired-end slam-seq?

[aleighbrown]

Rather than just sequencing the 3'utrs, what we've done is a standard paired-end RNA-seq with the addition of 4SU and the conversion step, I see that HISAT-3N should work fine for slam-seq, and pair-end bisulfate sequencing.

# Single-end slam-seq reads (T to C conversion, RNA) alignment with the standard 3N-index: hisat-3n -x genome -f -U read.fa -S alignment_result.sam --base-change T,C --no-repeat-index

But before I got down the rabbit hole of trying to use HISAT-3N, I just wanted to check if you think it would work okay

# Paired-end slam-seq reads (T to C conversion, RNA) alignment with the standard 3N-index: hisat-3n -x genome -f -1 read_1.fa -2 read_2.fa -S alignment_result.sam --base-change T,C --no-repeat-index

Because while on one strand of a paired-end read we have a T > C mismatch, and in the second read we would have A > G mismatch

Would I have to build 2 indexes and align separately?

[imzhangyun]

Hello Anna,

Thank you for using HISAT-3N. Yes, HISAT-3N supports paired-end slam-seq reads. HISAT-3N will handle the (T->C, A->G) problem internally. Just like the HISAT-3N supports paired-end BS-seq reads, it can align paired-end slam-seq reads without any problem. Please make sure you used the --base-change T,C for slam-seq alignment.

Let me know if you have any other question.

Leo