Hi,

Let's I have 6 samples - A, B, C and A', B', C'. Here A', B', C' are technical replicates of A, B and C respectively.

And A,B,C are biological replicates. So are A',B',C'. How do I map these samples using tophat2?

According to the website, it's best to map them separately if we want to do differential expression. And I plan to cuffdiff another set of samples (D,E,F) which are similar to A,B,C.

I would normally do something like this for biological replicates,

tophat2 -G annotation.gtf -o sampleA_results Bowtie2/genome  sampleA.fq
tophat2 -G annotation.gtf -o sampleB_results Bowtie2/genome  sampleB.fq
tophat2 -G annotation.gtf -o sampleC_results Bowtie2/genome  sampleC.fq

But with technical replicates, I'm not sure how to proceed. Should I combine the fastq files or should I just do the same command like above on A',B' and C' too? In the end, I plan doing cuffdiff. With bio replicates I can just do the following:

cuffdiff /refs/mm9/Annotation/Genes/genes.gtf -L G1,G2 A.bam,B.bam,C.bam D.bam,E.bam,F.bam

I'm not really sure how to proceed with technical replicates now in the picture. Any help on this would be appreciated.

I'm working on mouse data, single end reads.

Thanks!