Enhanced expression of exogenous genes from retroviruses using HIV2/Tat.
The capacity to produce large amounts of protein in mammalian cells is important in several contexts, including large-scale generation of biologically useful proteins, gene therapy, and transdominant genetics in cultured cells. For transdominant genetics, retroviral vectors are especially useful for delivery of expression libraries. However, even the potent CMV promoter is often unable to stimulate single-copy production of protein beyond the 1 microM level. We have adapted the HIV2/Tat expression system to retroviral vectors to boost expression above levels attainable with CMV promoters. We show that the system produces protein levels in four cell types tested which exceed levels attained by wild-type CMV or modified CMV promoters. In one cell line, the increase is 10-fold above CMV. Coupled with a stable expressed protein, levels of about 4 microM can be produced from presumptive single-copy retroviral transductants, and 30 microM from multicopy transductants.
Sandrock
Robert W
RW
Deltagen Proteomics Inc., 615 Arapeen Drive, Suite 300, Salt Lake City, UT 84108, USA. rsandrock@deltagenpro.com
Peterson
Isaac
I
Evans
Cameron
C
Wheatley
William
W
Kamb
Alexander
A
D001709:Biotechnology / Q000379:methods*; D016923:Cell Death; D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000235:genetics*; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D015498:HIV-2 / Q000235:genetics*; D007202:Indicators and Reagents / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000235:genetics; D012190:Retroviridae / Q000235:genetics; D054322:tat Gene Products, Human Immunodeficiency Virus
D015696:Gene Products, tat; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D049452:Green Fluorescent Proteins