any extra steps between using tophat and cufflinks

Question

[zizigolu]

Hi,

When I was using bowtie2, my advisor asked me to do some changes on my sam files to have a high quality sam. Now by tophat, I have some accepted_hits.bam files. Then if I should do something for my bam files before using them as an input for cufflinks?

Thank you

[GouthamAtla]

Filtering based on mapping quality is advised.

[Devon Ryan]

Wouldn't that muck with cufflinks' expression estimates?

[zizigolu]

thank you both

Answer 1

[Devon Ryan]

Cufflinks can get seriously slowed down by trying to assemble rRNA or repeat loci. It might considerably speed things up to try and filter those out (presumably bedtools intersect could be used for that).

Answer 2

[poisonAlien]

Cufflinks skips transcripts with high abundance and denotes its expression as 'high_fpkm' instead of numerical value. Change parameter --max-bundle-frags from its default value 500000 to higher value.