Hello everybody,
I have 120 sequence read files (76bp illumina reads) that I have converted into fastq format and wish to use in Tophat to align to the human reference genome and then view the alignments in the Integrated Genome Viewer (IGV).
I have access to a high powered computing facility so was going to submit by 120 fastq files seperately (so they can run in parallel) and have them output to seperate folders. If I do this, will I be able to simply concatenate the 3 main output files from each Tophat run (i.e. junctions.bed, coverage.wig and accepted_hits.sam) to gain 3 master output files for all 120 runs?!
As a test, before I run all 120 files, I have run a single fastq file. I have then used sam tools to:
Finally I have opened IGV and loaded hg18 before loading accepted_hits.sorted.bam. The viewer informs me that it has located the .bai file and automatically loaded it but then I see no alignments in the genome viewer I have tried zooming in on regions where, I believe, the .sam file is telling me there should be an alignment but I still see nothing.
Any help would be much appreciated. I have looked for 2 days to try and find the answer so I'm really sorry if I've missed a relevant post but I'm at my wit's end.
Cheers
I have 120 sequence read files (76bp illumina reads) that I have converted into fastq format and wish to use in Tophat to align to the human reference genome and then view the alignments in the Integrated Genome Viewer (IGV).
I have access to a high powered computing facility so was going to submit by 120 fastq files seperately (so they can run in parallel) and have them output to seperate folders. If I do this, will I be able to simply concatenate the 3 main output files from each Tophat run (i.e. junctions.bed, coverage.wig and accepted_hits.sam) to gain 3 master output files for all 120 runs?!
As a test, before I run all 120 files, I have run a single fastq file. I have then used sam tools to:
- Convert .sam to .bam [samtools view -bt hg18.fa.fai accepted_hits.sam > accepted_hits.bam]
- Sort my .bam file [samtools sort accepted_hits.bam accepted_hits.sorted]
- Create a bam index (.bai) file [samtools index accepted_hits.sorted.bam]
Finally I have opened IGV and loaded hg18 before loading accepted_hits.sorted.bam. The viewer informs me that it has located the .bai file and automatically loaded it but then I see no alignments in the genome viewer I have tried zooming in on regions where, I believe, the .sam file is telling me there should be an alignment but I still see nothing.
Any help would be much appreciated. I have looked for 2 days to try and find the answer so I'm really sorry if I've missed a relevant post but I'm at my wit's end.
Cheers
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