How to view and analyze BAM files from SOLiD Bioscope alignment

Question

[Aura]

Hello. I have some .bam, bam.bai and .sam files corresponding to a Bioscope mapping of genome reads from SOLiD to a reference genome. I have had several issues even openig the files. As far as I a go I was able to open the alignment in Samtools, but not in IGV. I have to say that I am new at this so probably I am doing this wrong. When I try to open the .bam file in the graphic interface by clicking Tools>Run igvtools...>Count, etc, I got a .tdf null file and in the terminal I got these warnings:

Chromosome: gi|448814763|ref|NC_000962.3| not found in .genome file. Skipping.
No features were found that matched chromosomes in genome...

What is happening here? How else can I visualize this alignment?

Thanks for the help.

[Istvan Albert]

most likely the chromosome names do not match.

Answer 1

[Martombo]

apparently the "chromosome" to which at least some of those reads were aligned to is called gi|448814763|ref|NC_000962.3|. if you only google NC_000962.3 you'll see that it is Mycobacterium tuberculosis H37Rv, complete genome. what you need to do is to get the fasta sequence of this genome (from ncbi). then create an index of it and use it as your reference.

[Aura]

Thanks so much. That is right. The H37Rv strain is my reference genome. So, I created the index form the fasta file with samtools:

samtools faidx H37Rv.fasta

And then used the H37Rv.fasta.fai to create a genome file in IGV. Yet when agregate the H37Rv.genome file as reference I dont see the grafic representation of it in IGV. Am I doing it right?

[Martombo]

Did you check that the header of the fasta sequence is exactly gi|448814763|ref|NC_000962.3|?

[Aura]

Yes I have, and It is. Also, I would like to get the annotation file for this genome (gff). It is a way to generate it from any of the ncbi formats?

[Martombo]

mmmhh... maybe something went wrong while producing the .genome files? I would simply load the indexed fasta sequence into IGV, with Genomes -> load genome from file. If that doesn't work I'm all out of ideas...

[Aura]

That worked! Thanks.