Total Bacterial Load via qPCR
Quantitative PCR (qPCR), also known as Real-Time PCR, is a method that measures the number of copies of a DNA region defined by a particular PCR primer(s). Using this method, we specifically amplify the 16S amplicon and quantify the total bacterial content in each sample to determine total bacterial load.
This technique can also be used as a means to determine the feasibility of your microbiome study by providing an exact measure of bacterial abundance in your samples. We highly recommend using this technique prior to sequencing samples suspected to have low biomass, to determine whether these samples will result in positive amplification.
DNA Extraction & Quality Control
DNA is extracted from samples using the Mag-Bind® technology and hand-held 96 channel electronic pipette. The use of magnetics beads for DNA extraction is fundamental for automation and to specifically capture DNA while excluding organic inhibitors. In-house testing has shown that the DNA extraction kit used has a good balance of DNA yield and quality, as demonstrated on a variety of environmental sample types.
qPCR Protocol
We use a 16S qPCR assay to determine 16rRNA gene copy number for each sample against our standard curve.
