The CRISPR/Cas9 method allows new genes to be inserted. After Cas9 cuts the Target-DNA, it can use a homologous piece of DNA as a donor template for homology-directed repair. But HDR only occurs when there are sticky ends, and Cas9's cuts end in blunt ends.

What did I miss?


1 Answer 1


According to this Addgene blog:

There are four different HDR pathways to repair DSBs. Here are three central steps of the HDR pathways:

  1. The 5’-ended DNA strand is resected at the break to create a 3’ overhang. This will serve as both a substrate for proteins required for strand invasion and a primer for DNA repair synthesis.

  2. ...

Essentially, this resection process generates the required overhangs through partial degradation of one of the strands.

To understand the mechanism of resection a bit better, this is a good read:

Huertas, Pablo. "DNA resection in eukaryotes: deciding how to fix the break." Nature structural & molecular biology 17.1 (2010): 11-16.


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