Cas13 system

I was wondering if anyone has successfully used the cas13 (LwaCas13a, PspCas13b, or RfxCas13d) system in mammalian cells for gene know down. What are your experiences with these systems?

let me know.


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Mint Lichover 4 years ago

One of my friends tried 13a and 13b, but does not seem to have KD. 13d is kind of having an effect, but far less effective than what they claimed in the paper. Probably need some optimization. Hope they have a protocol paper in the future.

Olive Kelpieover 4 years ago

Do you know specific details about how the Cas13a/b/d were delivered, and what genes were targeted? We find that the expression of the guide or protein can have an effect on the level of knockdown, so different delivery methods or cell types can have various efficiencies of knockdown.

Mint Lichover 4 years ago

I believe he used 293T cells and lipo2000. I am not sure how much plasmid is used.

White Elfover 4 years ago

It's likely that your friend will need to deliver more plasmid as the amount of Cas13 complex expressed in the cell needs to be high.

Here was a recent paper that came out that used RNP with great success. That might also be a good approach: https://www.cancerletters.info/article/S0304-3835(18)30381-1/fulltext.

This paper from Patrick Hsu also had good results with Cas13a/b and even better results with Cas13d: https://www.cell.com/cell/fulltext/S0092-8674(18)30207-1. It might be good to try different orthologs/systems to see what works

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