Comparing different method for DESEQ2
2
0
Entering edit mode
Vapin • 0
@dd8ababf
Last seen 8 weeks ago
Germany

Hi, this could be the general question regarding differential gene expression analysis.

Method 1: Mapping with STAR/HISAT2, counting the reads with HTSEQ, then DSEQ2.

Method 2: Count without alignment with Salmon and DESEQ2.

I usually do with the traditional method 1. But when I am comparing the two methods, getting about more than 60 percent differentially expressed genes? Did anyone experience that, or Did anyone do the same as I? What could be the potential reason for this difference?

DESeq2 GeneExpression • 292 views
ADD COMMENT
0
Entering edit mode

Are the same transcriptome annotations used for all the indexes and by HTSEQ? That could be a source of variance. Also, please clarify the difference - are you saying 60% of the significant genes are not shared between the two approaches? What are your cutoffs? Maybe plot the log2FCs of the two methods against each other.

ADD REPLY
1
Entering edit mode
@mikelove
Last seen 1 day ago
United States

You are performing DE across the same samples?

There are a number of differences, hence the development of methods like RSEM, kallisto, and Salmon, but I think most of all you would expect genes in gene families (with sequence homology) to be "up-regulated" in methods that don't throw out multi-mapping reads but proportionally assign them.

ADD COMMENT
0
Entering edit mode
swbarnes2 ★ 1.1k
@swbarnes2-14086
Last seen 1 hour ago
San Diego

HTSEq is way less smart about dealing with ambiguously aligning reads than salmon. I'd believe salmon. If you want to stick with your STAR alignment, you could use RSEM; that program is smart about dealing with ambiguous reads. You just have to configure STAR to export a transcripome-based sam file.

ADD COMMENT
1
Entering edit mode

Just a note: Salmon can also “ingest” txome aligned reads.

ADD REPLY

Login before adding your answer.

Traffic: 552 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6