Add enzyme to Radinitio

[Le Min Choo]

Hi Radinitio developers,

Could you consider adding the enzyme ApeKI to radinitio? I am trying to add reference genomes to my existing stacks database, and radinitio seems to be the most straightforward way to do this, apart from the fact that ApeKI is not yet one of the enzymes supported by the programme. 

Thank you for considering my request!

Best wishes

Le Min

[Angel Rivera-Colón]

Hi Le Min,

Thanks for checking out RADinitio.

Currently, the software does not offer support for enzymes with more than one restriction cutsite sequence (like the GCWGC for ApeKI). Given how restriction enzymes are implemented in RADinitio, adding this would require some time for development and testing, so I cannot guarantee it being available quickly.

In the meantime, could you explain the goals of your experiment a bit more? This might help with providing a quicker alternative than waiting for the RADinitio implementation.

Thanks,

Angel

[Le Min Choo]

Hi Angel,

Thanks for the reply!

I currently have a fasta file of a chromosomal level genome assembly which I'd like to incorporate into my existing ddRADseq stacks database.

From the fasta file, I'd like to do an in-silico restriction digest to generate restriction fragments, and from the restriction fragments simulate fastq reads files from them, and from there run stacks (denovo_map.pl etc) on these in-silico generated ddRADseq fastq reads with the rest of my real ddRADseq output.

I've tried ddRADseqtools (https://github.com/GGFHF/ddRADseqTools/tree/master/Package) which generates the restriction fragments, but at the next step it complains that some of the fragments are missing one of the restriction sites at the 3' end, although I've yet to troubleshoot this fully. I've also tried generating reads from the fasta file of restriction fragments (using NGSNGS https://github.com/RAHenriksen/NGSNGS) but it seems to be generating reads from all over the fragments (as in normal Illumina sequencing from libraries prepared with fragmented DNA) instead of the just the 150 bp from either end, which is what sequencing of restriction fragments involves.

If you have any suggestions on what else I can try, please let me know!

Thank you very much!

Best wishes

Le Min

[Angel Rivera-Colón]

Ok, I see. Yes, from what I see NGSNGS is for simulating whole-genome shotgun data, so it might not be the best for this purpose. I haven't checked the tool in a while, but what I remember ddRADseqTools should allow you to generate the ddRAD reads from the in silico digest, so it might be worth it to troubleshoot the results you already generated. I would start there.

I can start looking into the updates to RADinitio, but it might take some time to implement and test while I work on other more pressing things. Since you have already made progress with other software, my advice is to not wait for a new RADinitio release, since I cannot guarantee it anytime soon, sadly.

Thanks,

Angel