Hepatic DNA and RNA analysis of dogs A and B. Liver biopsies were taken from dogs A and B on days 420 and 210, respectively. DNA and RNA were isolated. One microgram of DNA was subject to PCR amplification for detection of HD and helper-virus genomes. RNA was reverse transcribed using a cFVIII-specific primer with a unique 25 nucleotide tag. Subsequent RS-PCR amplification enabled discriminate amplification of tagged cFVIII cDNA molecules. (1) 100-bp ladder, (2) HD PCR, (3) helper-virus PCR, (4) cFVIII RS-PCR, (5) HD PCR–negative control, (6) helper-virus PCR-negative control, and (7) cFVIII-PCR–negative control. Vector copies/cell was determined by Sybr green incorporation quantitative PCR. One-tenth microgram of liver-derived DNA was amplified, and the value 1 μg DNA = 105 liver cells28 was used to calculate the value for vector copies/cell shown here.
