Comparison of an In-House Whole HIV-1 Pol Genotyping Assay to Viroseq for Detection of HIV-1 Drug Resistance Mutations in Patients Failing cART in Botswana

Comparison of an In-House Whole HIV-1 Pol Genotyping Assay to Viroseq for Detection of HIV-1 Drug Resistance Mutations in Patients Failing cART in Botswana

Project Level: 
Email Address: 
tshepimokaleng@gmail.com
Phone Number: 
+267 7 370-7257

Mokaleng’s project looks at optimising the recently improved Long-Range HIV Genotyping for Analysis of HIV Drug Resistance and HIV Clustering in the Botswana population. In this project, they genotype 80% of the full length HIV genome using a long range genotype method - a modification of the method developed by Gal et al. This method enables the critical ability to detect drug resistance in subjects who are already on anti-retroviral therapy, by sequencing the entire HIV genome and locating drug-resistance mutations. The method uses proviral DNA and viral RNA as templates for amplification. The long range genotyping focuses on the 4 PCRs - the first PCR is the amplification of the large fragment taking almost the entire HIV genome (viral RNA) using RT-PCR (cDNA synthesis).  The large fragment is used as a 2nd round template for the last three amplicons. The amplicons 1 and 3 are sent to Sanger for direct sequencing while amplicon 2 is cloned before sequencing.