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GeneSeq HIV Technology
GeneSeq™ HIV utilizes advanced, proprietary technology for a highly accurate reading of the genotype of HIV. The GeneSeq HIV process employs reproducible and accurate sample preparation and nucleic acid sequencing methods. Data analysis is based on published literature, international expert recommendations, and in-house analysis of tens of thousands of paired genotypes and phenotypes.
Click on the links below to learn more about the technology behind GeneSeq HIV.
Performing the test —
Sample preparation
DNA amplified from viral RNA by reverse transcription-polymerase chain reaction (RT-PCR), using proprietary procedures and oligonucleotide primers that ensure reliable amplification of any sample with a viral load of 500 copies or more, serves as the starting material for the sequencing reactions.
Nucleic acid sequencing
Nucleic acid sequencing in GeneSeq HIV is based upon the dideoxynucleotide chain termination method of DNA sequencing (dideoxy sequencing)
- Purified RTV-DNA is combined with specific primers, deoxy-nucleotides, dye-terminator dideoxy nucleotides, and thermostable polymerase, and the resultant reaction mixtures are incubated in a thermal cycler
- During multiple cycle sequencing reactions, DNA polymerase synthesizes the complementary strand of a single-stranded DNA template by the sequential addition of one of four fluorescently labeled deoxyribonucleotides (dATP, dCTP, dGTP, dTTP)
- The reaction products are then separated by electrophoresis using the ABI PRISM® 3700 DNA Analyzer. The analyzer uses fluorescence to generate an image of the sample data from which the DNA sequence can be determined
Analyzing the data —
Sequence interpretations (interpretation algorithm)
Customized GeneSeq™ HIV software is used to analyze the sequence files.
- GeneSeq HIV software translates the DNA nucleotide sequence into an amino acid sequence, and then compares the patient's amino acid sequence to the sequence of a drug-sensitive ('wild-type') reference virus
- Variations in amino acid sequence (mutations) from the wild-type reference virus are identified and reported
- Based upon the mutations detected, an interpretation algorithm based on published literature, international expert recommendations, and in-house analysis of tens of thousands of paired genotypes and phenotypes, is used to predict drugs to which the patient's virus may exhibit reduced susceptibility
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