Can I use only one sgRNA for my CRISPR Knock in?

20 May 2023 1 10K Report

Hello,

I am trying to achieve the CRISPR KNOCK IN of HDR 1.7 kb in HEK293T will one sgRNA will be efficient to get a knock in or else should I use two sgRNAs?

Kristian Bruun Laursen

In short, yes! One sgRNA would be sufficient assuming it is a potent sgRNA (high editing frequency). Now with a 1.7 kb insert, the frequency is going to be very low, so it is advisable to include a selection marker (commonly antibiotic or fluorescent marker). Also, you could consider various HDR templates such as single- vs double-stranded, and modified terminal bases vs unmodified template.

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