Evaluation of easy, non-destructive methods of DNA extraction from minute insects

Abstract

Although it is important to identify species of pest and natural enemy insects in agriculture, morphological identification alone is often difficult. Because a combination of morphological identification and DNA barcoding is very important for identifying indiscriminable species, there is a need to develop non-destructive DNA extraction methods. Here, we compared three general methods of DNA extraction—the Chelex method, PrepMan® Ultra Reagent, and DNeasy® Blood and Tissue Kit—in the search for a rapid, simple, and non-destructive method of DNA extraction from pest and natural enemy specimens. Amplification of PCR production was achieved with the Chelex method and the DNeasy® Blood and Tissue Kit. However, not all specimens tested by the PrepMan® Ultra Reagent method yielded amplification products. Observation of the morphological features of dried Encarsia formosa specimens revealed that all heads and leg segments remained intact after DNA extraction by each of the three methods. Almost all samples treated with the PrepMan® Ultra Reagent method had large, discernible morphological changes, in particular, of the antennae and forewings, which are important features for identifying minute wasps. In contrast, the antennae and forewings of specimens on which we used the Chelex method suffered little damage. Use of the DNeasy® Blood and Tissue Kit method did not alter the forewing features, but strong wrinkles appeared in the antennae. We discuss the DNA extraction techniques from the perspective of these results.