Figure 1.

Genetic analysis of 2bF9 lentivirus (LV)-tranduced BMT recipients. (a) PCR analysis of the 2bF9 transgene. Genomic DNA was purified from peripheral white blood cells from primary (1°) and secondary (2°) 2bF9 LV-transduced hematopoietic stem cells recipients. FIX^null and C57BL/6 wild-type (WT) mice were used as controls. 2bF9 transgenic mouse DNA was used as a positive control for the 2bF9 transgene. Absence of the WT mFIX and presence of the disrupted mFIX gene confirmed that these mice were in the FIX^null background. Shown is one representative experiment that was performed three times. (b) Real-time quantitative PCR determined the average copy number of the 2bF9 transgene per cell in BMT recipients. Genomic DNA was purified from peripheral white blood cells, and 150 ng of DNA was analyzed for the 2bF9 transgene with normalization to ApoB. Bars represent mean ± SD. There were two groups of 2bF9 LV-transduced recipients. One was preconditioned with lethal total body irradiation, 1,100 cGy, the other with sublethal 660 cGy. For individual mice analyzed more than once over the course of study, the average copy number was calculated. Genomic DNA from FIX^null and 2bF9 homozygous transgenic mice was used as controls.