Question: GC content for RNA-Seq data
0
gravatar for sukesh1411
2.3 years ago by
sukesh141130
South Africa
sukesh141130 wrote:

Hi

I am new to the rna-seq data analysis. I have rna-seq data of leaf. I would like to perform transcriptome assembly using trinity. Before doing it, i removed the adapters and low quality bases. When i check the quality of reads in FASTQC it shows error for per base sequence content, per base GC content and sequence duplication levels. I dont know how to normalize this data before going to perform denovo assembly

Fastqc results

Thanks

rna-seq • 2.9k views
ADD COMMENTlink modified 2.3 years ago • written 2.3 years ago by sukesh141130
1

FastQC is probably overreacting again, doesn't look like something to worry about in my opinion.

ADD REPLYlink written 2.3 years ago by WouterDeCoster41k
4
gravatar for genomax
2.3 years ago by
genomax73k
United States
genomax73k wrote:

Don't do anything. That is very characteristic signature of RNAseq data in FastQC. If you were over cautious in removing low quality bases then dial back on that as well. You probably need to filter at Q20 or below if you truly have bad quality data.

ADD COMMENTlink modified 2.3 years ago • written 2.3 years ago by genomax73k

Hi genomax, just wondering, is it necessary to account for GC content when doing the differential expression analysis? Based on what it says on this article

ADD REPLYlink written 2.3 years ago by biofalconch380
1

In general, no. it's not. Because you are comparing the expression of gene X with gene X across samples, they will all have the same GC content.

If you would compare gene X (GC=35% ) with gene Y (GC=76%) then that would matter, but that's not the scope of differential expression analysis.

ADD REPLYlink written 2.3 years ago by WouterDeCoster41k

Thank you for your answer Wouter :)

ADD REPLYlink written 2.3 years ago by biofalconch380

I guess it depends whether there is a GC bias between different sequencing samples

ADD REPLYlink written 22 months ago by russhh4.7k
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