Genomic integration analysis.
To detect integrated vector DNA in the liver and spleen of treated mice, a nested PCR was performed with primers specific for the lentiviral vector and for mouse B2 family repeats. Adult SCID mice were injected with 109 TU/mL HIV-GFP lentiviral vectors, which were killed after 1 month (lanes 2,4,6) or 3 months (lanes 8,9). Mice injected with PBS served as control (lanes 1,3,5,7). The integrated lentiviral genomes in liver and spleen yielded a 121-bp and a 166-bp fragment at their 5′ end (left gel) and 3′ end (right gel), respectively. The top panel corresponds to the PCR products from the B2-vector PCR reaction. The middle panel corresponds to the nested PCR products with the use of the lentiviral vector-specific primers. The lower panel is from control nested PCR performed with 10 ng nonamplified genomic DNA from liver and spleen of treated and control mice. No DNA was added in the PCR reactions in lane C.
