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dCas9(D10A) and AncBE4max for Polynesian Albinism single nucleotide conversion
#1
Hello!

I'm working on a personal project to convert a Guanine into an Adenine in this specific albinism causing allele: https://www.ncbi.nlm.nih.gov/gene?Db=gen...;Term=4948

The mutation is at 2324 in the code.
My plan is to use a double nickase dCas9(D10A) to bring the AncBE4max to that site and convert the Guanine into Adenine while in-vivo.

I took inspiration from this article in particular: https://www.nature.com/articles/jhg2009130?proof=t

And this one: https://www.mdpi.com/2073-4409/9/7/1690/htm

As well as this article: https://www.nature.com/articles/s41467-019-11514-0

I haven't yet built a plasmid from scratch and am trying to use GenSmart Design to do so, but am not yet able to figure out how to delete things from the prebuilt plasmid sections. I am hoping to get some aide into what programs I should be using to build my plasmid as well as any know-how into how this works and if it can work.

Using the sgrna design tools online with the basic PAM of NGG, I haven't been able to find sites close enough to the edit zone to seem viable. I am trying to keep it below a span of 40 nucleotides, but I also don't have a good idea of what I'm doing. Any help with explaining how I should be going about this would be wonderful.

Due to certain personal circumstances, I am unable to continue my education at university and am trying to find other ways to continue my education into this field. Any tips for that would also be greatly appreciated.

Godspeed,

- Drig
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