We performed a CRISPR to knock out XPF in a RPE1 cell line and want to "pre-validate" clones through UV-C hypersensitivity (254 nm lamp). Which dose would you suggest to use to maximize differences in viability between knockouts and wildtype?
Many thanks in advance!
Victor
Hi Victor,
So, you're trying to kill the KO's with UVC to demonstrate that you've deleted XPF? If that's the case, I recommend setting up a "dose response" experiment with both the WT and KO's in parallel. I would say that you need at least 3 different doses to see the difference between the cell line: low dose, medium dose, and high dose.
If you have a non-adjustable lamp, figure out your energy output with a photometer, then calculate the time necessary based on that. Also, please consult the literature in addition. This will give you a better idea of where to start for this particular cell line. And also, make sure you have potential KO's and WT cells frozen down in dry-storage, just in case. I hope this helps.
-Bryan
Thank you Brian, we will be running a kinetics on the 2-20 J/m2 range, already found a significant effect on wt at 20 while according to literature 2 should be close to 90% proliferation.
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