I have generated a pileup of Illumina Miseq NGS data in IGV by mapping the reads onto the reference sequence reconstructed from the reads in order to check how well the sequencing has worked. The pileup image shows each base positions and also how many reads cover that position. I need information regarding how many reads have mapped, position of the reference where each read has mapped and also its pair (these are paired-end reads). How is it possible to extract this information in a text file? Also the coverage is not uniform with one end of the sequence having unusually higher coverage than the rest. Is there any explanation for this?
If you loaded that alignment as a BAM file, the BAM/SAM files already contain the kind of information you ask for. You could use samtools outside IGV or a combination with some one-liner scripts (such as https://gist.github.com/davfre/8596159) to get it calculated/printed out.
Bedtool will good tool for you. also samtools are recommended above.
Hi, other than your bioinformatics question that were answered above, your sequencing data looks somewhat suboptimal. What are you organism are you sequencing, what reference sequence did you use, how did you enrich for the sequence, and how did you make the libraries. This will help answer the last part of your question. Note that the information you are asking for is manageable if you are looking at a few exons or even a gene. In might become unwieldy quickly if you are looking at a lot data.
Shale
- Badges
- Science method