Transcriptomic assembly Stringtie

Question

[Nelislam4]

Hi,

I have replicates of RNA-seq sample from the same sample but on different lanes, and I need to run transcriptomic assembly by Stringtie so I have run their tutorial and merge the gtf files from every sample of replicates by StringTie --merge into a single gtf file that has the complete transcripts that were partially assemblies before merging. Then I have to use this merged.gtf file with every BAM file from the replicates in StringTie -eB to calculate their abundance.

Here is the point, the replicated Bam files definitely had duplication, so What can I do with those Bam files? I have to merge the bam files? but If I did so I will end up with duplication, and I knew that I shouldn't mark duplicate the reads.

so, What can I do to get the exact abundance from those Bam files against the merged gtf file "that was assemblies from the gtf files of those Bam files?

Please Help, Thanks in advance

[ATpoint]

Simply merge the BAM files prior to stringtie. You only make it overly complicated to keep them separated. They are the same poo, of RNA/cDNA so combine them.

[Nelislam4]

Thanks for your kind response, merging bam files prior to stringtie solve the issue of merging gtf files from each bam file; however, the coverage still an issue cos there will be replicates from the same sample......so How can I solve this problem??