Journal: PLoS Pathogens
Article Title: The Host Cell Sulfonation Pathway Contributes to Retroviral Infection at a Step Coincident with Provirus Establishment
Figure Lengend Snippet: The sulfonation pathway does not influence reverse transcription or the level of integrated virus DNA. (A and B) CHO-K1 cell lines were challenged with the VSV-G pseudotyped MLV vector LEGFP. Total DNA (A) or DNA from isolated nuclei (B) were harvested at 0 or 24 hpi, DNA concentration was quantitated by A260, and a real-time PCR amplification analysis was performed to measure the levels of viral DNA that were synthesized. (C) Untreated CHO-K1 cells or CHO-K1 cells that had been pretreated for 16 hrs with 100 mM chlorate and maintained in medium containing this concentration of chlorate were challenged with the MLV vector and subsequently analyzed for reverse transcription products as described in panel A. (D) CHO-K1 cells that were either untreated or treated with 100 mM chlorate, IM2 cells, and MCL7 cells, were challenged with the same VSV-G pseudotyped MLV vector and total DNA was harvested at 1 or 18 days post infection for real time PCR quantitation of viral DNA products. The chemically-mutagenized MCL7 cell line displays a strong block to MLV DNA integration  . Chlorate treated CHO-K1 cells were passaged in medium containing 100 mM chlorate for the duration of the experiment. The data shown in panels A–D are the average mean values obtained in independent experiments performed with triplicate samples and each is representative of three independent experiments. Error bars indicate the standard deviation of the data.
Article Snippet: DNA concentration was calculated by measuring the A260 on a SPECTRAmax Plus 96 well UV spectrophotometer (Molecular Devices, Sunnyvale, CA).
Techniques: Plasmid Preparation, Isolation, Concentration Assay, Real-time Polymerase Chain Reaction, Amplification, Synthesized, Infection, Quantitation Assay, Blocking Assay, Standard Deviation