I am working with paired-end 150 bp RNA-seq data and plan to use STAR followed by rMATS for alternative splicing analysis. The FastQC report is attached.
Based on this QC profile, what Trimmomatic parameters would you recommend?
Specifically, what values would you use for:
My goal is to preserve splice junction information and obtain the most reliable alternative splicing results with STAR and rMAT
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The data look mostly fine to me. Also, STAR’s not a global (end-to-end) aligner by default, so it has some tolerance for imperfect read ends.
So, I wouldn’t use aggressive settings for Trimmomatic. Unlike some other trimming programs, I don’t think Trimmomatic has default trimming values. But the program website gives example command/settings for PE and SE reads, so I’d start there. For example, LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36.
I'd also compare STAR alignment/QC metrics before and after trimming rather than assuming trimming improves the splicing analysis.
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