What is the best tool for doing Quality Control checks on my raw RNA-seq data?

Question

[anamaria]

Hello,

I just got my Illumina RNA sequencing experiment results in fastq format. What would be the best software to do QC on this data? Can you please share some good tutorials on this subject?

Thanks Ana

Answer 1

[adam.faranda]

I have been using FastQC from the Babraham institute. It is easy to use and gives fairly comprehensive summary statistics. Its output is compatible with multiqc. Multiqc can be used to aggregate several qc reports into a single unified document.

FastQC: https://www.bioinformatics.babraham.ac.uk/projects/fastqc/

MultiQC: https://multiqc.info/

[anamaria]

thank you so much, for the actual analysis, after QC is done, would you recommend doing this: 1.Mapping/Alignment (in HISAT2), 2. Assign aligned reads to genes (in HTseq), 3.Differential expression (in DESeq2)

[adam.faranda]

Hi Ana; that is more or less what I am doing. From what I can tell this pipeline is generally considered valid and well accepted. In some pipelines there is a trimming step after running quality control, before running alignments. I've found this to have relatively little impact on my data, but my reads are high quality to begin with. The Babraham institute also makes a tool called TrimGalore.

[anamaria]

Thank you so much for this very valuable feedback!