Reduced Representation Library construction for SNP discovery

[fastidiousme]

Hi,

I am preparing to go for SNP discovery in a plant genome by constructing reduced representation libraries. I already have quite a few published papers for reference. However, I would be grateful if anyone could suggest a working protocol for selection of a suitable Restriction Enzyme (meth-sensitive or meth-insensitive) for an entire genome and for constructing an RRL using the same RE.

Thank you in advance.

[uranotaenia]

RAD Wiki

Have you had a look at the RADSeq Wiki?

https://www.wiki.ed.ac.uk/display/RADSequencing/Home

There is a working protocol for library prep there and an Excel spreadsheet for selecting enzymes based on genome size, GC content and predicted number of enzyme cut sites.

[fastidiousme]

Thank you very much Uranotaenia! I will check this link out and let you know if it helps.

[ngs.analysis]

RRL sequence -unique SNP calling in unaligned regions.

Hi, we have a prepared library of RRL sequences for the an un-assembled genome (tetraploid). the region of interest is 250 bases long, sequenced 72 BP single end. We are using 1 of its parent genome as a partial reference. We also need to call the SNPs from the region which has not been aligned to our partial reference genome??? can some 1 suggest some tools that will help in the downstream analysis.