Storage and shipping of tissue samples for DNA analyses: A case study on earthworms

Nowadays, molecular analyses play an important role in studies of soil dwelling animals, for example in taxonomy, phylogeography or food web analyses. The quality of the DNA, used for later molecular analyses, is an important factor and depends on collection and preservation of samples prior to DNA extraction. Ideally, DNA samples are frozen immediately upon collection, but if samples are collected in the field, suitable preservation methods might be limited due to unavailability of resources or remote field sites. Moreover, shipping samples over long distances can cause loss of DNA quality e.g. by thawing or leaking of preservation liquid. In this study we use earthworms, a key organism in soil research, to compare three different DNA preservation methods - freezing at -20 °C, storing in 75% ethanol, and freeze drying. Samples were shipped from the United States of America to Austria. The DNA of the samples was extracted using two different extraction methods, peqGOLD™ and Chelex^® 100. The DNA amplification success was determined by amplifying four DNA fragments of different length. The PCR amplification success is significantly influenced by preservation method and extraction method and differed significantly depending on the length of the DNA fragment. Freeze drying samples was the best preservation method when samples were extracted using the silica based extraction method peqGOLD™. For samples that were extracted with Chelex^® 100, storage in ethanol was the best preservation method. However, the overall amplification success was significantly lower for the extraction procedure based on Chelex^® 100. The detection of the small DNA fragments was higher and independent from the extraction method, while the amplification success was significantly reduced for the longer DNA fragments. We recommend freeze drying of DNA samples, especially when they have to be shipped for longer distances. No special packaging or declaration is needed for freeze dried samples, and the risk of thawing is excluded. Storage of freeze dried samples also reduces costs because samples can be kept at room temperature in a desiccator. It should be noted, that the extraction methods showed significant differences in DNA amplification success. Thus, the extraction method should be taken into account when choosing the preservation method.