Effects of sample processing on the detection rate of environmental DNA from the Common Carp (Cyprinus carpio)

Environmental DNA methods have been used to monitor the presence of aquatic vertebrates in natural systems, although detection of DNA in the environment is sometimes a challenge. In this study, we evaluated the effect of sample processing on the detection of a species' environmental DNA in the water. Specifically, we examined whether freezing and then thawing water samples prior to analysis was an effective method of preserving them. The detection of Common Carp DNA was lower in samples that were frozen and thawed than in samples that were not, even though there was no difference in the DNA concentration, which was included with the DNA undetectable samples. In both types of samples, the DNA detection rate tended to be higher in a 2-μL volume of template DNA solution than in a 5-μL volume. DNA was detected in all non-frozen samples that were analyzed using a 2-μL template, both in three wells (three PCR replicate reactions per sample) with 40 PCR cycles and in eight wells with 55 cycles. The detection of Common Carp DNA in samples that were frozen and thawed was likely to increase through the use of the TaqMan Environmental Master Mix, which is used recently to efficiently release PCR inhibition. Our results suggest that environmental DNA detection is influenced by the processing of water samples after collection and by PCR reaction conditions. Use of non-frozen samples and a smaller DNA solution are recommended for detection of environmental DNA with quantitative PCR assays.