Lentiviral Vectors

1. What is Lentiviral Vectors? Lentiviral vectors can effectively integrate exogenous genes or exogenous shRNA into host chromosomes to achieve the effect of persistent expression of target sequences. In terms of infection ability, it can effectively infect neurons, hepatocytes, cardiomyocytes, tumor cells, endothelial cells, stem cells and other types of cells, so as to achieve a good effect of gene therapy.For some of the more difficult transfection of cells, such as primary cells, stem cells, not differentiated cells, etc., using slow virus vector, can greatly improve the transduction efficiency of shRNA gene or purposes, and integration of shRNA gene or purposes of greatly increases the chance to host cell genome, to compare the convenient realization of shRNA gene or purposes of long-term, stable expression. Lentiviral vector refers to a viral vector derived from human immunodeficiency virus-1 (iv-1). The lentiviral vector contains the genetic information needed for packaging, transfection and stable integration, and is the main component of the lentiviral vector system. Lentivirus vectors carrying exogenous genes, with the assistance of lentivirus packaging plasmids and cell lines, are packaged by viruses to become infectious virus particles. The expression of exogenous genes in cells or living tissues can be realized by infecting cells or living tissues. Lentivirus vectors contain the genetic information needed for packaging, transfection and stable integration. Lentivirus-packing plasmids provide all the helper proteins needed to transcribe and package RNA into recombinant pseudoviral vectors.To produce a high degree of virus particles, need to use expression vector and packaging plasmid transfection cells, at the same time for viruses in the cells of packaging, packaging good fake virus particles is secreted into the culture medium of extracellular, centrifugal made that after the supernatant fluid, can be directly used for host cell infection, after the purpose gene into the host cell, through the reverse transcription, integrated into the genome, and high levels of molecules expression effect.

2. Constructed by removing the cis-acting sequences needed for packaging, reverse transcription, and integration of the hiv-1 genome, it provides the proteins needed to produce viral particles in a trans-functional manner. The packaging components are usually constructed separately on two plasmids, one expressing Gag and Pol proteins and the other Env protein, with the aim of also reducing the likelihood of reverting to a wild-type virus. By co-transfected the packaging components with the three plasmids of the carrier components into the cells (such as human kidney 293T cells), the hiv-1 carrier particles with the target gene and only the ability of one-time infection without the ability of replication can be harvested from the superscript of the cells. The plasmid expressing vesicular stomatitis virus (VSV) glycoprotein G and the plasmid expressing dual tropic mouse Lentiviral vector envelope protein Env were used to replace the plasmid expressing HIV itself envelope protein Env, respectively, so that the hiv-1 vector particles were coated with VSV or dual tropic MLV envelope. The results of doing so have at least three positive significance: 1.the replacement of the envelope further reduces the possibility of the recovery of lentiviral vector into wild type virus. 2.the scope of HIV vector infection to the host is no longer limited to CD4+ cells, but expanded to almost all tissue source cells;The envelope of VSV endows the lentivirus carrier particles with a high degree of stability, enabling them to be concentrated through ultra-fast centrifugation and reach a high titer. Naldini. have achieved the titer of lentiviral vectors from 105 TU/ml to 108TU/ml.