Evaluation of methods of soil DNA extraction for PCR.

摘要

Different methods of direct DNA extraction and purification from soil samples were investigated on four soil types. The method using Sephacryl S-400 HR MicroSpin™ columns was the most robust to yield DNA suitable for PCR amplification using the fungal universal primers ITSI-F/ITS4. Some physical and chemical soil characteristics were determined and related to DNA yield and quality, and PCR amplification success. The A₂₆₀/A₂₃₀ and the A₂₆₀/A₂₈₀ ratios, and the PCR success, were significantly affected by the method of DNA extraction used.; The developed method was used to extract DNA from soil from five Populus tremuloides Michx. stands in an attempt to study the diversity of the endomycorrhizal fungi community. A nested PCR was performed using primers NS1/NS8 first, followed by primers VANS1/NS4. However, the first PCR usually gave weak or no amplification so it is suggested to determine the sensitivity of the method and improve PCR conditions for enhanced specificity.