CRISPR/Cas9 

The CRISPR/Cas9 system derived from bacterial adaptive immune systems is one of the most powerful genome editing technology. It is an RNA-guided genome editing tool that consists of a Cas9 nuclease and a single-guide RNA (sgRNA). By base-pairing with a DNA target sequence, the sgRNA enables Cas9 to recognize and cut a specific target DNA sequence, generating double strand breaks (DSBs) that trigger cell repair mechanisms and mutations at or near the DSBs sites. CRISPR/Cas9 technology has been studied extensively and its application has been expanded from the modification of the gene in cells to organisms. The potential role of CRISPR/Cas9 in gene therapy has made it to become one of the hottest pots in cancer treatment. Different concepts of CRISPR/Cas9-mediated cancer therapy, including tumor-related genes manipulating, tumor immunotherapy, tumor research modelling and anti-cancer drug resistance overcoming are established in various cancer types.

The greatest advantages of the CRISPR-Cas9 system are its simplicity and wide applicability in genome manipulations of almost all biological systems tested to date, including cell lines, stem cells, yeasts, worms, insects, rodents, and mammals. For a targetable DNA site, only a corresponding 20 nucleotide gRNA is needed to guide the CRISPR-Cas9 to cut the target DNA at the desired location. The repair of the broken DNA ends occurs either through NHEJ to generate indels, which has been used to generate random genomic mutations or through HDR in the presence of donor oligonucleotides or DNA fragments containing homologous sequences flanking the DSB sites to generate precise site-directed nucleotide or large gene replacements, leading to generation of targeted gene mutations or corrections.