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Methods HIV-1 Subtype C molecular infectious clone pIndieC (Genebank AB023804) was used to construct two experimental vectors, pIndieC consInC RTWILD and pIndieC consInC RTSYNM, which are identical except for the presence of four synonymous nucleotides differences in Integrase. Standard curve was prepared for each vector using linear plasmid DNA template. The K103N, V106A, V106M, Y181C resistance mutations were placed in the pIndieC consInC RTWILD and replication fitness of each mutant was compared with that of wild type in a head to head growth competition assay. Virus supernatant harvested at day 0, 4, 8, 12 for RealTime quantitative PCR assessment. Relative fitness was estimated using online tool http://bis.urmc.rochester.edu/vFitness and significant differences in relative fitness between wild and mutant were determined with Mann Whitney test. |
