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Guidelines for the Qualitative Detection of Viral  Form

Guidelines for the Qualitative Detection of Viral Form

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[Music] in this video methods are demonstrated for quantifying and sequencing single hiv genomes from individuals with viral loads of less than 50 copies per milliliter of plasma first a tube containing seven milliliters of plasma is centrifuged to pellet the virus rna is then extracted to quantify the level of hiv-1 a single copy assay is performed in which the rna is reverse transcribed loaded onto a 96 well plate with standards and controls and subjected to quantitative real-time pcr the level of virenia is then calculated based on the input volume of plasma as few as 0.3 copies per milliliter can be detected to characterize the viral population a single genome sequencing assay is performed in which cdna transcribed from viral rna is spread over a 96-well plate to reach an endpoint dilution then nested pcr is performed to amplify viral genomes if 30 or fewer of the pcr reactions are positive for a given sample amplification of a single copy of cdna is indicated those samples are the

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