Análise comparativa da estabilidade do DNA de Dalbulus maidis (DeLong & Wolcott) (hemiptera: cicadellidae) sob diferentes métodos de preservação para uso em RAPD-PCR

Abstract

The appropriate preservation of the DNA of a certain organism is important for successful application of molecular techniques like RAPD-PCR. This study was designed to compare simple methods of preservation of specimens of Dalbulus maidis (DeLong & Wolcott) (Hemiptera: Cicadellidae) with respect to quantity and quality of DNA for use in RAPD-PCR, after different storage periods. Eight methods were tested: freezing (-20oC); ethyl alcohol 70% (-20oC and room temperature); absolute ethyl alcohol (-20oC and room temperature); air-dry; and preservation in extraction buffer (whole and homogenized insect). At intervals of 10 to 30 days, insect DNA was extracted, quantified and amplified through RAPD-PCR using primer OPA-04. The quality of extracted DNA was observed on 0.8% agarose gel. After 210 days of preservation, freezing (-20oC) showed to be the best method. Satisfactory quantities of DNA were also obtained from insects conserved in absolute ethyl alcohol (-20oC), ethyl alcohol 70% (-20oC) and extraction buffer (whole and homogenized insect). Insects conserved in absolute ethyl alcohol (room temperature), ethyl alcohol 70% (room temperature) and air-dry conditions were inappropriate for RAPD-PCR studies after 120, 60 and 10 days of storage, respectively.