CMV-SpCas9 Lentivirus

Lentivirus and retrovirus belong to the family of Retroviridae. Lentivirus is named because it has a latent period of several years after infection. Compared with other retroviral vectors, lentiviral vectors can infect cells in both the dividing and non-dividing phases. Their complex genome structure gives lentiviral vectors a unique integration mechanism and persistence of infection. The most in-depth research is the lentiviral vector based on HIV-1 modification. It includes an expression plasmid and multiple auxiliary plasmids. It is usually used to co-transfect 293T cells with multiple plasmids to achieve virus packaging, and then the packaged recombinant virus is used to infect cells to randomly and stably insert the target gene into the host cell genome to achieve rapid and stable expression of the target gene.

As a common gene knock-in method, lentiviral vectors are often used to construct steady-state cell lines for gene expression. In addition, it is also used to mediate CRISPR/CAS9 technology. By constructing a lentiviral vector containing sgRNA sequences, the recombinant vector and lentiviral packaging plasmid are co-transfected into 293T cells. Collect viral particles to infect the target cell line, and obtain a stable co-transfected cell line through screening. Inside the cell, Cas9 and sgRNA combine to form a cleavage complex, which is transported into the cell nucleus to complete the cleavage of the sgRNA complementary DNA fragment (target gene), thereby achieving the purpose of gene knockout/knockdown.