Question: RNA read alignment using transcriptome
0
gravatar for evelyn
4 months ago by
evelyn100
evelyn100 wrote:

Hello,

I want to align RNA reads to indexed reference transcriptome. But I could not find a way to make a transcriptome reference index and align using that with hisat2. Is the command line same for reference genome or transcriptome for hisat2.

Thank you!

sequencing • 222 views
ADD COMMENTlink modified 4 months ago by ATpoint36k • written 4 months ago by evelyn100
0
gravatar for h.mon
4 months ago by
h.mon30k
Brazil
h.mon30k wrote:

The command should be the same as for building a reference genome index. However, as far as I know, hisat2 hasn't been widely used for this, the commonly used alternatives being bwa or bowtie2.

ADD COMMENTlink written 4 months ago by h.mon30k
0
gravatar for ATpoint
4 months ago by
ATpoint36k
Germany
ATpoint36k wrote:

Yes, the indexing is the same but you do not provide a splice site file as the transcriptome (at least this is typically the case) already contains the spliced transcripts. There is specialized software for transcriptome quantification, such as salmon and kallisto, just that you know.

ADD COMMENTlink written 4 months ago by ATpoint36k

yes, I want to use salmon for quantification. But couldn't find specifically if I can use hisat2 for indexing the transcriptome and use it for alignment to create a bam file which can be used for salmon alignment mode. I am using:

hisat2-build transcriptome.fa transcriptome
hisat2 -q -x transcriptome -1 input_1.fastq -2 input_2.fastq -S output.sam
samtools view -bS output.sam > output.bam
salmon quant -t transcripts.fa -l -A -a output.bam -o salmon_quant

Is this approach okay. Thank you!

ADD REPLYlink written 4 months ago by evelyn100

This post might help:

A: Mapping onto the Transcriptome, RNA-seq

ADD REPLYlink written 4 months ago by Fatima590

You do not need any alignment prior to using salmon. It will happily accept fastq files. Please read its documentation https://salmon.readthedocs.io/en/latest/salmon.html#quantifying-in-mapping-based-mode

ADD REPLYlink written 4 months ago by ATpoint36k
Please log in to add an answer.

Help
Access

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.
Powered by Biostar version 2.3.0
Traffic: 827 users visited in the last hour