DNA methylation and quantitative analysis of B19V nucleic acids in transfection experiments.

UT7/EpoS1 cells were transfected with 1 µg/106 cells of either unmethylated or M.SssI methylated B19V DNA, excised from pB19-M20 plasmid, and samples collected at 2, 24, 48, 72 hours post-transfection (hpt). Quantitative analysis was carried out for the determination of the amounts of viral DNA and total viral RNA from cell fractions (copies/104 cells). Methylation index was calculated by parallel amplification with both MSP primer pairs on bisulfite-modified DNA and is expressed as ratio of methylated over total viral DNA.