Marlowe N, Hoo B, Dileanis J, Elbeik T; International Conference on AIDS.
Int Conf AIDS. 2000 Jul 9-14; 13: abstract no. TuPeA3009.
N. Marlowe, Pe Biosystems, 850 Lincoln Centre Drive, Foster City, CA 94404, United States, Tel.: +1 650 554 2328, Fax: +1 650 638 6177, E-mail: marlownm@pebio.com
Background: Increasing prevalence of non-subtype B strains in the AIDS pandemic requires detection of drug resistant mutations in all HIV-1 subtypes. Objective: To assess the performance of PE Biosystems Version 2 HIV Genotyping System (HGS v2) on well-characterized HIV standards from the Walter Reed subtype panel representing 30 HIV-1 isolates of subtypes A through G. Methods: HIV-1 subtype stocks were diluted to 25,000, 5,000 and 1,000 viral particles/ml and viral load were determined using the Bayer Quantiplex HIV-1 bDNA assay. HIV-1 RNA was isolated from the 30 subtype isolates using HGS v2 Sample Preparation module. RT-PCR amplification and subsequent bi-directional sequencing of the 1.8 Kb amplicons was performed with HGS v2 RT-PCR and Sequencing modules. Sequencing data were collected on ABI Prism 377 DNA Sequencer and analyzed using PE Biosystems Sequencing Analysis v3.3 and HGS v2.1 software. Results: 30/30 (100%) samples at 25,000 viral particles/ml were successfully amplified. Region of pol encompassing 1-99 codons of protease (pro) and 1-315 codons of reverse transcriptase (rt) gene was sequenced and analyzed. Sequence comparison of pro and rt gene of HIV-1 subtypes genotypes to reference strain NL4-3 indicates a range of homology and allows detection of the mutations in the positions known to be associated with drug resistance in subtype B isolates. To determine the sensitivity of HGS v2 7/7 (100%) isolates representing subtypes A, B, C, E, F, were amplified and detected down to 1000 viral particles/ml. Viral genotypes were identical from different dilutions of the same viral stock. Conclusion: The PE Biosystems HGS v2 can be used for drug resistance analysis for group M HIV-1 subtypes with sensitivity at 1,000 viral particles/ml. Studies are in a progress to evaluate the sensitivity of the assay on all HIV-1 subtypes.
Publication Types:
Keywords:
- Acquired Immunodeficiency Syndrome
- Branched DNA Signal Amplification Assay
- Drug Resistance
- Drug Resistance, Viral
- Evaluation Studies
- Genes, pol
- Genotype
- HIV Infections
- HIV Seropositivity
- HIV-1
- Research Design
- Reverse Transcriptase Polymerase Chain Reaction
- Substance Abuse Detection
- Viral Load
- genetics
Other ID:
UI: 102238855
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