Figure 4.

Semiquantitative analysis of hFVIII retroviral vector (GCsamF8EN) DNA and RNA sequences in recipients at 26–35 wk posttransplant. Genomic DNA and total RNA were prepared in parallel from the same BM source. Representative data shown are from 8 of 13 analyzed GCsamF8EN BM transplant recipients. (A) Vector DNA was measured via coamplification of hFVIII vector sequences and mouse β-globin sequences by semiquantitative PCR, using NIH 3T3/GCsamF8EN DNA quantitative standards (Left). The estimated percent gene transfer for experimental animals (1–8) is shown at the top of each lane (Right). T, tolerant. NT, nontolerant. (B) Vector-specific hFVIII RNA was measured by semiquantitative RT-PCR using Raji/GCsamF8EN (Raji/FVIII) RNA and BM RNA from LNL6 recipients as positive and negative controls respectively. RT, reverse transcriptase. β₂-m, β₂-microglobulin. (−), PCR reactions without added reverse transcriptase; (+), reverse transcriptase added.