Two simple techniques for the safe Sarcoptes collection and individual mite DNA extraction.
Availability of mites is a recognized limiting factor of biological and genetic investigations of the genus Sarcoptes. Current methods of deoxyribonucleic acid (DNA) extraction from individual mites also need substantial improvement in efficiency and operator friendliness. We have first developed a technique for efficient and safe extraction of living mites from scabietic skin samples (crusts or deep skin scrapings). Its core device is a large plastic syringe connected with a 1.5-ml Eppendorf tube. The source material is introduced in the syringe and the device in a shoe box with the tip half of the tube emerging. Mites migrate towards a heat source during a minimum of 36 h. Then, the tube is detached and the mites utilized without risks for the operators. A second technique allows operator-friendly manipulation of individual mites for DNA extraction. Fixed mites are isolated by adhesion to a small strip of polyvinyl chloride (PVC) adhesive tape operated with tweezers. Then, mite and strip are plunged in the lyses buffer and the sample twice submitted to thermal shock for disruption of the chitinous exoskeleton. Data show that the tape does not interfere with successive DNA extraction with a commercial kit. The corresponding protocol, that we briefly name "PVC adhesive tape + thermal shock + kit DNA extraction," compares favorably with the available ones.
Soglia
Dominga
D
Dipartimento di Produzioni Animali, Epidemiologia ed Ecologia, Università degli Studi di Torino, 10095 Grugliasco, Italy. dominga.soglia@unito.it
Rambozzi
Luisa
L
Maione
Sandra
S
Spalenza
Veronica
V
Sartore
Stefano
S
Alasaad
Samer
S
Sacchi
Paola
P
Rossi
Luca
L
D000818:Animals; D004247:DNA / Q000302:isolation & purification*; D008967:Molecular Biology / Q000379:methods*; D010274:Parasitology / Q000379:methods*; D040021:Sarcoptidae / Q000235:genetics*
D004247:DNA