This week we finally got around to talking about the recent CRISPR/Cas9 papers in Drosophila. We mostly focused on Bassett et al, but also discussed Gratz et al.
Like other Drosophila researchers we’ve talked to, we are excited by these techniques and we can’t wait to try them out in the lab (Our new postdoc, Byoungchun Lee will be doing a lot of the work here) – CRISPR/Cas9-mediated mutagenesis will very likely develop into a “must-have” genetic technique for model organism labs.
Questions/comments we had: We mostly talked about how we would use this technique for our lab, with our favorite genes. In the Bassett et al paper the efficiency and frequency of mutagenesis for y and w looked good (although there was a high level of lethality – even in the non-injected controls ?!). But will this high efficiency hold true for all genes? Will genomic/chromatin context matter? (some of our favorite genes are in heterochromatic regions) Also, what’s the most efficient fly injecting/crossing scheme to generate stable mutant stocks (especially if the mutation efficiency ends up being quite low for some genes)? Finally, we’re interested to see how well the combination of CRISPR/Cas9 with donor oligos can be used to engineer specific alterations in our favourite genes. Gratz et al did a great job of using the approach to introduce attP sites into yellow, and presumably a similar approach can be used to introduce specific nucleotide substitutions into our favourite genes (e.g. to generate serine-to-alanine mutations at known phosphorylation sites).
Overall, two great papers. Exciting times ahead for fly genetics!
2 thoughts on “This week’s lab journal club: CRISPR/Cas9 mutagenesis in Drosophila”
How concerned are you about linked, off-target mutations here?
Hi Dave – thanks for the comment. Off-target mutations should be no problem, even if they are tightly-linked with the targeted mutation – I think we just have to treat CRISPR-induced mutants like any other (P-element, EMS) and rely on standard fly genetics to validate any mutant phenotypes (e.g using rescue constructs, multiple alleles, recombine-out any 2nd hits). The CRISPR approach will just be more versatile and MUCH quicker than current mutagenesis approaches.
Looking forward to hearing about some of your lab journal clubs….
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