CRISPR (Clustered regularly interspaced short palindromic repeats) loci and their Cas (CRISPR associated) genes encode an adaptive immune system that protects bacteria and archaea from viral (phage) and plasmid infection. The RNA-guided DNA endonuclease Cas9 associates with the dual RNA guides (CRISPR RNA [crRNA]) and trans-activating RNA [tracrRNA]) or a synthetic single-guide RNA (sgRNA) and cleaves double-stranded DNA targets complementary to the guide RNA. The double-stranded break occurs 3 bp upstream of the PAM site, allowing targeted sequence modifications via two DNA repair pathways: non-homologous end joining (NHEJ) that introduces frame shift insertion and deletion (indel) mutations leading to loss-of-function alleles and homology-directed repair (HDR) for precise insertion of point mutations or a fragment of desired sequence at the targeted locus. At present, the CRISPR-Cas9 system has been applied to a variety of model organisms, including human, mice, rats, zebrafish, elegans, plants and bacteria.
Advantages of CRISPR Technology
• The target can be designed simply and quickly, without having to construct endonuclease repeatedly.
• RNA-guided genomic DNA specific recognition needs no consideration of DNA methylation.
• Targeting operation can be highly efficient and flexible. Mutiple genes can be edited at the same time and genetic modification can be genetic.
• CRISPR has the same or highly efficiency of gene editing comparing ZFN and TALEN.
• Knock-out and knock-in cell lines can be established.
• CRISPR/Cas9 lentivirus system can infect division and non-division cells.
• CRISPR can be applied to any species.
Genome editing refers to an emerging branch of biotechnology that is earlier genetic engineering technologies. By using these biotechnologies researchers are able to target specific DNA sequences and induce a double stranded break, taking advantage of recombination to create synthetic genetic genome of a host.