After finishing STAR two-step alignment, I got 62% uniquely mapped reads. But featureCounts gives me only 17% successfully aligned rate.
featureCounts -T 8 -F GTF -p --countReadPairs -t exon -g gene_id -a ~/genome_ref/gencode.v38.annotation.gtf -o ~/expression/all_counts.txt *.bam
I also have a look at other data in this dataset. Many of them are around 20% successfully aligned rate. But QC report is fine. According to the library preparation protocol, the library is unstranded. I also checked the bam file in IGV. The reads distribution seems to be normal. (Although I don't know how to get an overview of the read peaks referring to this answer)
Previously when I analyze another dataset, this rate could be around 70%.
I have 2 questions:
Does low aligned rate severely affect the quantification of gene expression? In another word, can these data be used for downstream analysis?
Why this happens? Any solutions or explanation?