Counting reads (not coverage...)

[smol]

Hi
Hopefully someone knows of some software or a script that will save me having to write one….

I want to put together some summary stats for my RAD sequence data (70bp Illumina single reads with a restriction site at one end). The reads are spread throughout the genome depending on the presence of the restriction site, and stack up directly on one another rather than forming contigs. I want to get a count of the number of reads for each particular sequence, so that I can get a frequency distribution of read coverage. I’ve tried using GATK DepthofCoverage walker, but this gives coverage per base relative to the reference, so it gives the sum of two sequence’s coverage where the sequences overlap.

My data is in SAM format, but I can’t simply do a count on chromosome position because the reverse strand sequences are 70bp away from the forward sequences.

Any ideas?

Thanks
Sam

[kmcarr]

Have you looked at this software - Stacks. I came across it a while back when I was investigating RAD sequencing for a researcher. The project never happened so I never used the software hence I can't say for sure if it will do what you need but it's worth a look.

[smol]

Thanks kmcarr, Stacks is a great software, but it would mean repeating my analysis from scratch...

Anyway, I have realised that I was wrong about the coordinates in my SAM file - they are all given from the same position, so I can count the reads from the SAM file.

Problem solved!

[kmcarr]

... but it would mean repeating my analysis from scratch...

That's why it's called research, not just search!

Glad you worked it out.

[Ekeberg]

Hi Smol, hope you can check this mail soon))
I want to do exactly the same, tutorials like the ones for ggplot don't help me so much. Can you please let me know how you constructed your graphic.
I appreciate your advice, tks!