Monday, November 15, 2004
D0015

Moving toward Thysanopteran phylogeny: DNA extraction methods that preserve voucher specimens

Cheryle O'Donnell, caodonnell@ucdavis.edu, Diane E Ullman, and Michael P. Parrella, mpparrella@ucdavis.edu. University of California-Davis, Department of Entomology, One Shields Avenue, Davis, CA

Our ultimate goal is to estimate thrips phylogeny using morphological and molecular characteristics. Reliable thrips identification using morphological methods requires use of a compound microscope to view cleared specimens mounted on slides. For molecular phylogenetic research, it is essential that specimens from which DNA is extracted are identified accurately. However, preparation for molecular analysis is usually destructive, and identification based on other thrips from the same collection may be confounded by the presence of more than one species in a sample. To avoid such misidentifications, we developed a technique that preserves the thrips exoskeleton for mounting as a morphological voucher, while extracting strong DNA product for our molecular analysis. We make a small incision along the abdomen, and place the whole thrips into a QiagenTM DNeasy tissue kit, proteinase K lysis buffer (ATL) for processing. We do not grind up the specimen. The thrips exoskeleton is then retrieved from extraction kit, dehydrated and mounted in Canada balsam on a microscope slide. We also extracted thrips DNA using the methods of Sunnucks and Hales (1996) and D.C. Morris (pers.com.), but obtained lower DNA yields than with our method.


Species 1: Thysanoptera Thripidae Frankliniella occidentalis (western flower thrips)
Species 2: Thysanoptera Thripidae Heliothrips haemorrhoidalis (greenhouse thrips)
Species 3: Thysanoptera Thripidae Thrips palmi
Keywords: molecular phylogentetics, voucher specimen