How to best use Stacks

[Loic Pittet]

Dear all,

I have been using Stacks for many years, but still I am wondering what would be the best option to process my new data. I would be happy to hear your opinion on that. Here is what my dataset is made of:

• 100bp Single-end RAD seq reads
• 75 samples representing three species (two diploid parents, one tetraploid)
• No references for any species

Should I build a denovo reference using only the parent species and then map the tetraploid to it... Is it actually possible to do so ? I would really appreciate your advices !

Best,

Loïc

[Catchen, Julian]

Hi Loïc,

 

What you decide to do would depend on how the three species are related to one another, and how you expect the RAD loci to be shared among them. But, you can experiment with different setups. Without knowing the details of your system, I would probably try constructing three different de novo datasets: 1) Parent/species 1 catalog then matching the tetraploids to that catalog; 2) Parent/species 2 catalog and the tetraploids matched to it; 3) Maybe all three together in one catalog – it depends again on how close they all are to one another. I would also optimize my assembly parameters independently for the three species before I decided on a final set of parameters. Then, you would want to see how the tetraploids map to the different parental species, but again, it depends on what you are hoping to show. A final alternative would be to select the best de novo dataset (that is, the one where the most total loci can be mapped to the catalog), then turn that into a reference and align the other species to it doing a reference-based analysis.

 

Best,

Julian