Visualizing Bowtie Output In Genome Browser

Question

[k.nirmalraman]

I received a text file which has following entries and I assume this is a Bowtie aligned map files and I would like to visualize the same on a local genome browser.

HWI-ST863:202:C2014ACXX:4:1101:2197:1894 1:N:0:NTTTTTCG + NM_011363 3354 TGTTGTTGTTGTTTTAAACAAAATGGAAAAGCATAA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 1

HWI-ST863:202:C2014ACXX:4:1101:3133:1962 1:N:0:NAATAAGA + NM_175190 729 GTGTGGTTGTCCCTTTTGTTAATAAACATATGAGCA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 0

HWI-ST863:202:C2014ACXX:4:1101:5688:1889 1:N:0:NTATCTGG + NM_020606 4378 CTGTCTTGATCCATTTCTTCTGCATGATTCCAGAAA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 0

HWI-ST863:202:C2014ACXX:4:1101:9703:1907 1:N:0:NTTAAAGG + NM_026030 397 GCTTCTGATGACTTAGATGATTTGAACTTCTTTAAT IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 0

I would like to validate if the reads are enriched towards the 3' end of the mRNA (genes). I would also like a local genome browser like tool for this purpose. How can I see this in a genome browser like IGV?

[k.nirmalraman]

I guess I should some how convert this file to bed format...

[Jordan]

I don't quite understand the question. But if you just want to visualize the bam file, you can load the BAM file in IGV browser. And you can load your reference genome as well and just focus on the region you are interested in.

[k.nirmalraman]

I tried to load this file directly into IGV from file-> load file with mouse genome as reference, but I cannot see anything beyond that

[Jordan]

Does your BAM file have an index file? I think IGV usually needs it.

[Nicolas Rosewick]

You also need to have the same chromosome name between the bam and your reference sequence

[k.nirmalraman]

I don't have a index file, but here it says, I should have a sorted file. I tried IGV tools for sorting but no succes. File type or sorting not supported error... which leaves me wondering, if the file is in bam format....

[Jordan]

you can use samtools for sorting the bam file.

samtools sort file.bam file.sorted

then, index the bam file (Note: to index the bam file, the file needs to be sorted beforehand).

samtools index file.sorted.bam

Answer 1

[Alex Reynolds]

Convert your sequences to FASTA and run them through, for example, a command-line instance of UCSC BLAT to get a PSL file. Run the PSL file through BEDOPS psl2bed to get a BED file. You can turn this into a BED-like BedGraph file pretty easily for uploading to and visualization with a, for example, local UCSC Genome Browser instance.

Answer 2

[swbarnes2]

For starters, can you convert that to .bam format? (Or to .sam, which can then be converted to .bam)

Bams don't have '+' to indicate the strand, it's in the flag value instead.