| Abstrakt: |
ABSTRACTTo obtain high-titer recombinant retroviruses, we constructed plasmid pDL, which carries the extended Psi region and the polyomavirus early region, including the replication origin and the early gene. Although pDLis useful for obtaining high-titer recombinant retroviruses, this vector plasmid is difficult to modify further for tissue-specific expression of foreign genes. To overcome this problem, the coding region of the polyomavirus early gene was expressed in the packaging cell lines. We modified the packaging cell lines, 2 and PA317, by stably introducing the polyomavirus early gene, and established MP34 and MP37 from 2, and PAMP51 from PA317. In the transient expression system using plasmids with and without the polyomavirus replication origin, the titers of recombinant retrovirus produced by these cell lines were 10100 times higher than produced by the parent cell line and reached levels of 0.51.5 106cfu/ml. Expression of the polyomavirus early gene in the packaging cell lines did not stimulate the production of replication-competent retrovirus. We also routinely established stable clones producing retrovirus titers over 1 107cfu/ml from MP34 and PAMP51. We found that the activity of the long terminal repeat (LTR) promoter is stimulated by the polyomavirus early region protein(s) in these cell lines. Therefore, increases titer can be expected to occur in all the retroviral vectors in which LTR promoter is used to transcribe the retroviral genome. |
