Question: Mapping query sequence with RNA-Seq
0
gravatar for skevalkumar
3 months ago by
skevalkumar0 wrote:

I am trying to map query sequence with RNA-Seq data. I have tried with BWA-MEM and HISAT2 that gives many reads. However, it does not provide any read with 100% match with query read. I appreciate recommendations in terms of software and parameters.

rna-seq alignment • 182 views
ADD COMMENTlink modified 3 months ago • written 3 months ago by skevalkumar0

I am trying to map query sequence with RNA-Seq data. I have tried with BWA-MEM and HISAT2 that gives many reads. However, it does not provide any read with 100% match with query read.

Let me try to put it a slightly different way and you can confirm is this is correct.

You have RNAseq data i.e. your query(ies). When you try to align those reads (presumably to a reference genome or transcriptome) you are not able to get any alignment?

ADD REPLYlink written 3 months ago by genomax60k

No, I have my query sequence (around 4kb) and trying to align with one of my RNA-Seq data. When I am getting reads, there is not a 100% match of my read with my query sequence.

ADD REPLYlink written 3 months ago by skevalkumar0
1

You should do these alignments the other way. Use your RNAseq data as the query against your reference (4 kb) and see what happens.

ADD REPLYlink written 3 months ago by genomax60k

As in less than 100% align or none of the reads align perfectly?

ADD REPLYlink written 3 months ago by Devon Ryan87k

Have you eyeballed your RNASeq data? Isn't your 4kb sequence far longer than any of your reads? How can you expect them to match the query 100% along its whole length like that?

ADD REPLYlink written 3 months ago by swbarnes24.7k

None of the reads aligned perfectly with query sequence

ADD REPLYlink written 3 months ago by skevalkumar0
1

Try doing the mapping by flipping your search. Use the 4kb sequence as reference when you search with your reads.

ADD REPLYlink modified 3 months ago • written 3 months ago by genomax60k
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