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Table 1 Modified Cas9 approaches to increasing the frequency of homology-dependent repair outcomes

From: Strategies to improve homology-based repair outcomes following CRISPR-based gene editing in mosquitoes: lessons in how to keep any repair disruptions local

Approach Rationale Reported effect References
Cas9-i53 (hypothetical) Expression of i53 alone strongly suppressed the recruitment and function of 53BP1 at DSBs Cas9-i53 fusion has not been reported, however expression of i53 in cell lines induces BRCA1 accumulation at DSB sites similar to 53BP1 knockout cells Canny et al. [28]
Cas9-DN1S DN1S inhibits the recruitment of key NHEJ proteins such as RIF1 two- to threefold increase in HDR and three- to fourfold reduction in NHEJ Jayavaradhan et al. [24]
Cas9-yRAD52 yRAD52 can stimulate strand invasion and homologous recombination at a DSB site Threefold HDR increase in human cells
Threefold HDR increase in chicken cells
Shao et al. [44], Wang et al. [45]
Cas9-HE (CtIP) CtIP assists in end-resection and promotes HDR Twofold HDR increase in human cell lines, human iPSCs, and rat zygotes Charpentier et al. [49]
Cas9-UL12 UL12 recruits endogenous MRN complex which resects DSBs and promotes HDR Twofold HDR increase in HEK293 cells Reuven et al. [51]
miCas9 (cas9-Brex27) Brex27 recruits RAD51, a pro-HDR protein that searches for homologous sequences and facilitates D-loop formation at DSB sites Up to threefold HDR increase depending on donor size in human cell lines Ma et al. [56]
REDIT Exploits bacteriophage SSAPs to insert kilobases of sequence at DSB sites Threefold increase in HDR in human AECs and iPSCs Wang et al. [61]
Cas9-RecA RecA is able to promote single-strand annealing pathway at DSBs in between repetitive sequences 2.5-fold increase in SSA in HEK293T cells Lin et al. [66]
Cas9-hGem Restricts Cas9 presence and activity to late S and G2 when HDR occurs most frequently 1.87-fold HDR increase in HEK293T cells Gutschner et al. [79]
Cas9-PCV Ensures proximity of donor DNA at DSB site through PCV-ssDNA covalent bonding Up to 30-fold increase in HDR correcting frameshift mutations with short ssDNA donor Aird et al. [84]