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22marks t1_iy1z5m4 wrote

Reply to comment by Smeghead333 in How exactly does CRISPR-CAS9 insert new genes? by AutomaticAd1918

Are there just random sequences always floating around for repairs? Forgive the analogy from a layperson, but is there any form of checksum? How does it know it needs to grab that sequence, especially since it’s not even the original sequence it’s replacing? Simply because the ends match like perfect “puzzle pieces” and it’s like “good enough?”

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Smeghead333 t1_iy20oib wrote

Normally, when a break happens, there's another copy of the DNA sequence in the cell - remember you have two copies of each chromosome: one from your mom and one from your dad. So the repair mechanism looks for another similar sequence and copies it (oversimplifying here) to patch the hole.

With CRISPR, if you inject a few thousand or million copies of the altered sequence you want, the odds are very good that the repair system will grab one of those instead of the non-altered sequence on the other chromosome.

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CrateDane t1_iy4yv1k wrote

>Normally, when a break happens, there's another copy of the DNA sequence in the cell - remember you have two copies of each chromosome: one from your mom and one from your dad. So the repair mechanism looks for another similar sequence and copies it (oversimplifying here) to patch the hole.

HDR is mainly active in S and G2 phase, where you get up to four copies of each chromosome - two maternal, two paternal. That provides additional templates for repair (or let's say a stalled replication fork ripped both paternal sister chromatids apart - you then still have two maternal templates available).

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