dCas9(D10A) and AncBE4max for Polynesian Albinism single nucleotide conversion - Printable Version
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dCas9(D10A) and AncBE4max for Polynesian Albinism single nucleotide conversion - Drig - 06-30-2021
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=gene&Cmd=DetailsSearch&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.