title: DNA recovery from microhymenoptera using six non-destructive methodologies with considerations for subsequent preparation of museum slides.
abstract: Because of the tiny size of microhymenoptera, successful morphological identification typically requires specific mounting protocols that require time, skills, and experience. Molecular taxonomic identification is an alternative, but many DNA extraction protocols call for maceration of the whole specimen, which is not compatible with preserving museum vouchers. Thus, non-destructive DNA isolation methods are attractive alternatives for obtaining DNA without damaging sample individuals. However, their performance needs to be assessed in microhymenopterans. We evaluated six non-destructive methods: (A) DNeasy® Blood & Tissue Kit; (B) DNeasy® Blood & Tissue Kit, modified; (C) Protocol with CaCl2buffer; (D) Protocol with CaCl2buffer, modified; (E) HotSHOT; and (F) Direct PCR. The performance of each DNA extraction method was tested across several microhymenopteran species by attempting to amplify the mitochondrial gene COI from insect specimens of varying ages: 1 day, 4 months, 3 years, 12 years, and 23 years. Methods B and D allowed COI amplification in all insects, while methods A, C, and E were successful in DNA amplification from insects up to 12 years old. Method F, the fastest, was useful in insects up to 4 months old. Finally, we adapted permanent slide preparation in Canada balsam for every technique. The results reported allow for combining morphological and molecular methodologies for taxonomic studies.
authors.affiliation: a Universidad Autónoma de Nuevo León, Av. Universidad s/n. San Nicolás de los Garza N.L. México 66451.
authors.forename: Adriana J
authors.initials: AJ
authors.lastname: Guzmán-Larralde
authors.affiliation: b Centro Nacional de Referencia de Control Biológico, Dirección General de Sanidad Vegetal, SAGARPA. Km 1.5 Carretera a Tecomán Estación del FFCC Tecomán, Colima, México 28120.
authors.forename: Alba P
authors.initials: AP
authors.lastname: Suaste-Dzul
authors.affiliation: b Centro Nacional de Referencia de Control Biológico, Dirección General de Sanidad Vegetal, SAGARPA. Km 1.5 Carretera a Tecomán Estación del FFCC Tecomán, Colima, México 28120.
authors.forename: Adrien
authors.initials: A
authors.lastname: Gallou
authors.affiliation: c INIFAP Campo Experimental General Terán, Carretera Montemorelos-China km 31, Nuevo León, México 67400.
authors.forename: Kenzy I
authors.initials: KI
authors.lastname: Peña-Carrillo
mesh_terms: D000818:Animals; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification*; D003576:Electron Transport Complex IV / Q000235:genetics; D007313:Insecta / Q000145:classification* / Q000235:genetics*; D009144:Museums; D017422:Sequence Analysis, DNA
chemical_list: D004247:DNA; D003576:Electron Transport Complex IV
keywords: COI; DNA extraction; extraction d’ADN; hyménoptères parasites; museum insects; parasitic hymenoptera; spécimens d’insectes de musée