Tophat/cufflinks with multiple fastq files?

[xhuister]

Hello,

I have multiple fastq files. I want to use tophat for the alignment and use cufflink to get the annotation. Because the memery limit, I need to process one file each time.

My question is:
do I need to merge the SAM/BAM files of tophat before using cufflink (1), or can I use cufflink for each SAM/BAM file and then use cuffmerge to merge the annotation files (2) ?

(1)
tophat file1.fq --> cufflink file1.sam
tophat file2.fq --> cufflink file2.sam
cuffmerge file1.gtf file2.gtf

(2)
tophat file1.fq
tophat file2.fq
sammerge file1.sam file2.sam
cufflink merge.sam

Thank you very much!

[dpryan]

Are the fastq files from the same sample?

[xhuister]

These fastq files are downloaded from NCBI. They can be considered as from the same sample.

Are the fastq files from the same sample?

[dpryan]

If they're from the same sample then they need to be merged either before or immediately after alignment. If you're looking for novel splice junctions, then you'll get better results merging before alignment.

[xhuister]

If they're from the same sample then they need to be merged either before or immediately after alignment. If you're looking for novel splice junctions, then you'll get better results merging before alignment.

Thank you dpryan. But I thought that merging before tophat or after tophat should have the same results. Why merging fq files before tophat can get better results?