Hello all,
I'm trying to make GBS libraries for damselflies. I'm using Pstl enzyme and everything looks good, but when I analyze the library in the bioanalyzer, there is a strange pike at ~200pb, at the beginning I thought that could be adapter dimers, but the size didn't fixe. Someone has had the same problem? What do you think it is?
Thaks a lot,
Is there a chance that the damselfly genome contains a highly repetitive sequence bearing a PstI site?
Would say highly repetitive region as well. I would test other enzymes and see if you get more even size distribution of fragments...
I ran a simulated digest on the Ladona fulva genome and didn't find any RAD fragments overlapping with repeats around 200bp, but this of course doesn't necessarily mean that there isn't a highly repetitive region in your case. Results attached (tabulated as counts of fragments at each possible fragment size)- note that the Missing_sites column are counts fragments that are NOT flanked on one or more ends by a restriction enzyme (e.g. are terminated by contig/scaffold ends). If there happen to be any more closely related genome, you can run the script on that
Scripts here: https://github.com/tkchafin/fragmatic
One enzyme: fragmatic.pl -r "CTGCA^G" -i
Or with two enzymes: fragmatic.pl -r "CTGCA^G C^CGG" -i
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