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Non-homologous end joining (NHEJ) is a pathway that repairs double-strand breaks in DNA. It is called "non-homologous" because the break ends are directly ligated without the need for a homologous template, in contrast to homology directed repair (HDR), which requires a homologous sequence to guide repair.
[111] [112] Small molecules can also be used to improve homology directed repair, [113] often by inhibiting the non-homologous end joining pathway and/or the theta-mediated end-joining pathway. [ 114 ] [ 115 ] A system with the Cpf1 effector protein was created that is induced by small molecules VE-822 and AZD-7762. [ 116 ]
Microhomology-mediated end joining (MMEJ), also known as alt-non-homologous end joining, is another pathway to repair DSBs. The process of MMEJ can be summarized in five steps: the 5' to 3' cutting of DNA ends, annealing of microhomology, removing heterologous flaps, and ligation and synthesis of gap filling DNA. [5]
dsDNA-break repair pathways and genome editing using CRISPR-Cas nucleases. A common form of genome editing relies on the concept of DNA double stranded break (DSB) repair mechanics. There are two major pathways that repair DSB; non-homologous end joining (NHEJ) and homology directed repair (HDR). NHEJ uses a variety of enzymes to directly join ...
Cas9 (or "CRISPR-associated protein 9") is an enzyme that uses CRISPR sequences as a guide to recognize and open up specific strands of DNA that are complementary to the CRISPR sequence. Cas9 enzymes together with CRISPR sequences form the basis of a technology known as CRISPR-Cas9 that can be used to edit genes within living organisms.
The process of V(D)J recombination is mediated by VDJ recombinase, which is a diverse collection of enzymes. The key enzymes involved are recombination activating genes 1 and 2 (RAG), terminal deoxynucleotidyl transferase (TdT), and Artemis nuclease, a member of the ubiquitous non-homologous end joining (NHEJ) pathway for DNA repair. [4]
The SSA pathway is unique in that it does not require a separate similar or identical molecule of DNA, like the DSBR or SDSA pathways of homologous recombination. Instead, the SSA pathway only requires a single DNA duplex, and uses the repeat sequences as the identical sequences that homologous recombination needs for repair.
Other strategies include in planta gene targeting, whereby the homology repair template is embedded within the plant genome and then liberated using CRISPR cutting; [43] upregulation of genes involved in the homologous recombination pathway; downregulation of the competing Non-Homologous-End-Joining pathway; [39] increasing copy numbers of the ...