Environmental samples often contain inhibitors such as polyphenolics, humic/fulvic acids, tannins, melanins, etc. that affect downstream applications such as PCR. The Zymo-Spin III-HRC is designed to remove these PCR inhibitors and does not bind DNA. The Zymo-Spin III-HRC can be purchased separately as the OneStep PCR Inhibitor Kit (D6030).

Check sample type and input amount. Low biomass samples such as swabs, some soil samples, water, air are expected to have low yield. High biomass samples such as feces may cause overloading of the spin column. Using less input may improve yields.

Ensure sample is fully homogenized. Depending on bead beater used, the settings may have to be optimized for each sample. The homogenization time may be extended to see if yields improve.

Apply heated elution buffer (60-70° C) and allow elution buffer to incubate on the column for several minutes prior to elution.

Ensure that the DNA Elution Buffer is added directly to the column matrix.

Low A260/230 values can be due to handling issues (e.g. transfer of column during wash steps). To minimize reagent wash buffer carryover, centrifuge the column at max speed for 1 minute prior to elution.

Spin down lysate and Binding Buffer mixture at max speed for 5 minutes, this will eliminate any debris not caught by Spin III-F filter. Also avoid debris in lysis tube when transferring lysate, spin at max speed after bashing for 1-3 minutes.

We have validated our kits with both high-speed homogenizers and low-speed disruptors. We highly recommend users to optimize their bead beating conditions for their own instruments. We recommend using a 2 ml-tube adapter to ensure that the bead beating is efficient (do not use adapters made of foam). You can reference the Optimized Lysis Protocols table under documents for some recommendations.

This is to remove any debris from the lysate that could interfere with the DNA binding to the silica membrane of the column.