I have human RNASeq data generated using lexogen kit. I have followed their analysis protocol (to process raw data). Then used tophat2 for alignment, observed only 53%. check unalinged using blast they are going human rRNA sequences end2end. Then i used hisat2, the % increased to 75. Then tried bowtie2 end2end parameter (which does not align junction read i assume) has given 79%.
Unaligned reads from different aligners checked against NCBI NT using BLAST they are hitting to human rRNAs/ncRNAs. Questions,
How bowtie is preforming better than junction aligners?
How do i improve the overall alignment?
Reference used was GRCh38 from ensemble
I have seen the same trend for fungi sample as well.
Any in site what could be the reason??
Did you use equivalent settings for each aligner? Could you post the command-line for each aligner?
used default parameters, bowtie2 end-end, same in tophat enabled b2-sensitive.