Per base sequence content error in RNAseq analysis

Question

[ahmad.sajad4541]

i am new to rna-seq analysis. i have downloaded paired end data from SRA database and upon checking these data through FASTQC it is showing me error on Per Base Sequence Content and Sequence Duplication Levels. The rest are all green except two samples are giving yellow flag for Per Tile Sequence Quality. I don't know what to do with this. is it normal for RNA-seq data for having these?

Secondly, the adapter contamination shows green flag, should i have to download adapter sequence? because i don't find any adapter sequence information in SRA website. Is it ok to use illumina universal adapter "TruSeq-3 "?

[GenoMax]

Please check: https://sequencing.qcfail.com/articles/positional-sequence-bias-in-random-primed-libraries/ for why you see that pattern

Answer 1

[ATpoint]

That's normal in RNA-seq and has no further implications. Actually, the only metric I look at in fastqc these days is adapter content which is fine here. Proceed as is. There are previous threads on fastqc+ RNA-seq, please find via a search engine.

[ahmad.sajad4541]

@ATpoint thank you for your response. Should i go for alignment step without trimming(using trimmomatics)?

[ATpoint]

Yes, I see no need for any trimming here.

[ahmad.sajad4541]

thank you so much