FIG. 5.

Production of reverse transcription intermediates in resistant cell lines. (A) DNA was harvested from infected and uninfected cells (5 × 10⁵/well of a six-well plate) of the indicated cell lines. QPCR was performed using a primer/probe set that recognizes the first-strand transfer (+SSS) reverse transcription intermediate of the MLV genome that was used in the initial screen. (B to E) Cells (5 × 10⁵/well of a six-well plate) of the indicated cell lines were infected with 5 × 10⁵ IU of LEGFP[VSV-G] at 4°C for 2 h. Total DNA was harvested at 0 hpi and 24 hpi. DNA concentration was quantitated by A260, and QPCR was performed using primer/probe sets that recognize the (B) minus-strand strong stop, (C) minus strand, (D) first-strand transfer (+SSS) products, and (E) second-strand transfer (plus strand) reverse transcription intermediates. The numbers of DNA molecules were determined by comparison to a standard curve generated from serial dilutions of plasmid DNA. The data shown are the average mean values obtained in an experiment with triplicate samples and are representative of results of three independent experiments. Error bars indicate the standard deviations of the data.