454 FLX Titanium fragment sequencing of 16S rRNA Lung HIV Microbiome sample CP4_V1V3_76482_BALSIDEA.sff
This protocol uses the creation of a pooled, barcoded amplicon 454 library using a set of barcoded primer pairs.  The V13 and V35 primers were designed at the Broad Institute and used in the Human Microbiome Project’s 16S survey of healthy subjects. Documentation: Quant-iT ds DNA Assay protocol by manufacturer’s specifications, SybrGreen Assay protocol by manufacturer’s specifications, poolingCalculator.xls, and MinElute PCR Purification Kit Manual. Method: The PCR will be carried out using AccuPrime Taq High Fidelity.  It is not necessary to setup this reaction on ice, however it is recommended. 