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Amplifying and Quantifying HIV-1 RNA in HIV Infected Individuals with Viral Loads Below the Limit of Detection by Standard Clinical Assays
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Chapters
- 00:05Title
- 02:00Extraction of RNA from Large Volumes of Plasma
- 04:58The Single Copy Assay
- 08:42Single Genome Sequencing
- 11:24Representative Single Copy Assay and Single Genome Sequencing Results
- 13:27Conclusion
Quantifying levels of HIV-1 RNA in plasma and sequencing single HIV-1 genomes from individuals with viral loads below the limit of detection (50-75 copies/ml) is difficult. Here we describe how to extract and quantify plasma viral RNA using a real time PCR assay that reliably measures HIV-1 RNA down to 0.3 copies/ml and how to amplify viral genomes by single genome sequencing, from samples with very low viral loads.
Tags
AmplifyingQuantifyingHIV-1 RNAViral LoadsLimit Of DetectionStandard Clinical AssaysViral GenesInsightViral DynamicsVirus Host InteractionsPatientsNatural ControlCombination Antiretroviral Therapy (cART)Single Genome Sequencing (SGS Protocol)Single-copy Assay (SCA Protocol)Real-time PCR AssaySensitivityVolume Of PlasmaRNA Copy NumberInternal Control TestingDNA ContaminationTriplicate MeasurementsSingle-genome Sequencing Assay (SGS)Non-labor IntensiveLimiting Dilution AssayEndpoint Diluted CDNA Product
