DNA Quantity and Quality in Remnants of Traffic-Killed Specimens of an Endangered Longhorn Beetle: A Comparison of Different Methods.
The sampling of living insects should be avoided in highly endangered species when the sampling would further increase the risk of population extinction. Nonlethal sampling (wing clips or leg removals) can be an alternative to obtain DNA of individuals for population genetic studies. However, nonlethal sampling may not be possible for all insect species. We examined whether remnants of traffic-killed specimens of the endangered and protected flightless longhorn beetle Iberodorcadion fuliginator (L., 1758) can be used as a resource for population genetic analyses. Using insect fragments of traffic-killed specimens collected over 15 yr, we determined the most efficient DNA extraction method in relation to the state of the specimens (crushed, fragment, or intact), preservation (dried, airtight, or in ethanol), storage duration, and weight of the sample by assessing the quantity and quality of genomic DNA. A modified cetyltrimethyl ammonium bromide method provided the highest recovery rate of genomic DNA and the largest yield and highest quality of DNA. We further used traffic-killed specimens to evaluate two DNA amplification techniques (quantitative polymerase chain reaction [qPCR] and microsatellites). Both qPCR and microsatellites revealed successful DNA amplification in all degraded specimens or beetle fragments examined. However, relative qPCR concentration and peak height of microsatellites were affected by the state of specimen and storage duration but not by specimen weight. Our investigation demonstrates that degraded remnants of traffic-killed beetle specimens can serve as a source of high-quality genomic DNA, which allows to address conservation genetic issues.
Section of Conservation Biology, Department of Environmental Sciences, University of Basel, St. Johanns-Vorstadt 10, 4056 Basel, Switzerland hans-peter.rusterholz@unibas.ch.
Hans-Peter
HP
Rusterholz
Section of Conservation Biology, Department of Environmental Sciences, University of Basel, St. Johanns-Vorstadt 10, 4056 Basel, Switzerland.
Sylvain
S
Ursenbacher
Natural History Museum, Augustinergasse 2, 4001 Basel, Switzerland.
Armin
A
Coray
Museum zu Allerheiligen, Klosterstrasse 16, 8200 Schaffhausen, Switzerland.
Urs
U
Weibel
Section of Conservation Biology, Department of Environmental Sciences, University of Basel, St. Johanns-Vorstadt 10, 4056 Basel, Switzerland.
Bruno
B
Baur
D000818:Animals; D001517:Coleoptera / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000032:analysis*; D056727:Endangered Species; D005858:Germany; D018895:Microsatellite Repeats; D021141:Nucleic Acid Amplification Techniques; D016133:Polymerase Chain Reaction; D013048:Specimen Handling / Q000379:methods*; D013053:Spectrophotometry / Q000379:methods
D004272:DNA, Mitochondrial
CTAB method; DNA quantity and quality; Iberodorcadion fuliginator; microsatellite; qPCR