Journal:

Article Title: Alpha Interferon Inhibits Hepatitis C Virus Replication in Primary Human Hepatocytes Infected In Vitro

doi: 10.1128/JVI.76.16.8189-8199.2002

Figure Lengend Snippet: Characteristics of the strand-specific HCV RNA assays used in this study. (a) Strand specificity of the positive-strand-specific HCV RNA rTth RT-PCR assay. Decreasing amounts of positive-strand (+) HCV RNA (100, 10, 1, and 0.1 fg) and of negative-strand (-) HCV RNA (10, 1, and 0.1 pg) synthesized from an appropriate plasmid were subjected to the rTth RT-PCR assay. The products were analyzed by agarose gel electrophoresis. (b) Strand specificity of the negative-strand-specific HCV RNA rTth RT-PCR assay. Decreasing amounts of negative-strand HCV RNA (100, 10, 1, and 0.1 fg) and positive-strand HCV RNA (10, 1, and 0.1 pg) synthesized from the same plasmid as for panel a were analyzed by the same procedure. (c) Range of linear quantification of the quantitative assay based on real-time PCR using the LightCycler technology and SYBR green I dye for detection. The range of linear quantification of the assay was studied by testing 10-fold serial dilutions of synthetic positive- and negative-sense HCV RNA strands after RT at 70°C with the rTth polymerase. Each point is the mean of three experimental values for each dilution. y is the slope of the linear plots.

Article Snippet: PCR products were purified with Microcon 100 before sequencing (Millipore, Dublin, Ireland).

Techniques: Reverse Transcription Polymerase Chain Reaction, Synthesized, Plasmid Preparation, Agarose Gel Electrophoresis, Real-time Polymerase Chain Reaction, SYBR Green Assay