What are the packaging steps for slow Lentivirus

Lentivirus is a retrovirus, it can target genes into some difficult to transfection of cells, such as primary cell, and target genes were randomly into the host genome, which greatly increased the transfection efficiency, and can be stable expression in cell lines, several generations can be stable cell line selection.
Because it is a random integration, there are uncertainties, there are some company provides fixed-point integration technology, target genes of fixed-point integrated into specific parts of the genome, and to ensure the efficient expression of cell does not produce random integration possible damage.

Lentivirus expression vectors contain the genetic information needed for packaging, transfection, and stabilization.
The lentivirus packaging plasmid provides all of the helper proteins needed to transcribe and package RNA to 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 used directly to the host cell.

Lentivirus vector is normal chain RNA genome, its genome into the cell, was itself to carry in the cytoplasm of reverse transcriptase reverse transcription of DNA, DNA to integrate former complex formation, after entering the nucleus, DNA integrated into the cell genome.
The integrated DNA transcription mRNA is returned to the cell pulp to express the target protein or generate RNAi interference.

The lentivirus packaging system consists of a packaged-plasmid mixture (Mix) and a LentiviralVector (LentiviralVector), as shown in the following figure (from MIT) :

The carrier contains the basic components of HIV 5 'ltr and 3 'ltr and other auxiliary elements.

Different packaging plasmids of different systems are different. For example, the three-plasmid system in this laboratory is used as an example. The packaging plasmid mixture contains pMDL, VSVG, prsv-rev, and the ratio is 5:3:2.
Including pMDL coding HIV are the main structural protein of gag gene and coding virus specific enzyme of pol, pRSV - Rev containing encoding regulation gag genes and gene expression regulation of pol factor Rev, VSVG contains provide source of herpes simplex virus virus packaging need VSVG genes.

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