Hi Ravi,

I got around this problem by using the fastq interlacer to join reads in to a single file, then use deinterlacer to output only reads that have a pair in the correct order.

You may need to alter read IDs first by adding /1 and /2 to the end (see interlacer help text). I used unix command line sed but I'm sure you can use galaxy tools to do this.

Shaun


Quoting Ravi Karra <ravi.ka...@gmail.com> on Wed, 22 Feb 2012 12:29:18 -0500:

Hello,

I have Illumina 76bp paired end data for a zebrafish RNA-seq experiment and am basically stuck while trying to pre-process my data prior to using Tophat/CuffDiff.

For each sample, I have a read1 fastq file and a paired read2 fastq file. After using FASTQ Groomer, I trimmed the ends using FASTQ quality trimmer with a threshold quality score of 20 ans a window size of 1 (I think that will essentially lop off the end of the read until the quality score is >= 20). Next, I trimmed the adapters using Clip.

What I am left with is a modified read1 fastq file and a modified read2 file, where the pairs are not in the same order and some reads are left without pairs. From what I have read, I don't think TopHat can incorporate paired end data that is out of order.. I tried to get around the ordering issue using FASTQ joiner, but this tool is not able to join the reads (return is 0 joined reads). I am not really sure why FASTQ joiner didn't work for me and am looking for suggestions of what to try next.

Thanks!
ravi
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