Using IGV to View Alignments

[fk566938]

Hello. I have some Bam files of illumina RNA-seq data aligned to the human genome that I would like to upload into the Broad's Integrative Genome Viewer. By doing this and putting in coordinates of a gene isoform of interest is it possible to:

1. View expression levels of the gene isoform per each bam file of particular tissue.

2. Be able to see the alignment (The base pairing) to be sure that the sequence we want is actually aligning with the RNAseq data.

[blancha]

1. Yes. The only caveat is that if you have several overlapping isoforms, it can be difficult to determine to which isoform the reads belong. Mind you, any software program you use will have the same problem, so you'll understand better how the isoform counts can sometimes be unreliable.

2. Yes, any mutations relative to the reference will be highlighted, and you can view the sequence of the reference genome, as well as the sequences of the aligned reads.

The BAM files have to be sorted and indexed for viewing in IGV.

[Michael.Ante]

Hi fk566938,
I'd like to add a technical remark:
Once you have your expression values (which will be most likely estimated read counts or FPKM values; like blancha mentioned), you can create a bed file with the isoforms' coordinates (e.g start position, the transcripts' centre), name, and expression. Then, you can include this bed file into the IGV session.

Cheers,
Michael