Quantitative PCR for bacterial and fungal abundances

As a proxy for bacterial and fungal abundances, 16S rRNA (bacteria) and ITS (fungi) gene copy abundances were quantified from soil DNA samples using Femto™ Bacterial DNA quantification kit (Zymo Research) and Femto™ Fungal DNA quantification kit (Zymo Research) following the manufacturer’s protocol. DNA extracts were diluted 1:10 prior to quantification and 1 µl of the diluted samples was used for each qPCR reaction. All samples were analyzed in triplicate. No-template negative controls were included in each run. Bacterial and fungal DNA standards were provided in the kit and the ng DNA standard per well was converted to copy numbers which were used for final calculations. qPCR reactions were performed in a CFX Connect Real-Time PCR Detection System (BioRad). qPCR efficiencies averaged around 85% and 90% for bacterial and fungal assays, respectively.