Avaliação de quatro protocolos de extração de DNAs em amostras teciduais cervicais infectadas pelo papilomavírus humano

Abstract

The Human Papillomavirus (HPV) is considered a sexually transmitted infection, being a determining factor for the development of cervical cancer. The prevention methodology of cervical cancer recommended by the Brazilian Unified Health System is the analysis of cytological alterations, popularly known as Papanicolau. However, the molecular diagnosis has been indicated as a technique more sensitive to the virus presence, which may be inactive in the host cells not necessarily causing an infection, yet having an impact in the prevention of the cancerous changes. One of the molecular techniques capable of detecting HPV is the Polymerase Chain Reaction (PCR). To do so, it is necessary to first extract the DNA from the collected sample, which impacts on the sensitivity of the diagnostic test. In order to standardize the methodology for better performance in tissue fragments, we compared four protocols for total DNA extraction in cervical tissue biopsies collected in women showing cytological alterations compatible with HPV infection in the city of Dourados-MS, Brazil. The following protocols were tested: 1) HotSHOT; 2) Wizard® Genomic DNA Purification Kit; 3) Boom; 4) Dnaesy Blood and Tissue Qiagen. The extracted samples were measured by spectrophotometry and amplified by the Polymerase Chain Reaction using the primers GH20 / PCO4 and Mucosal HPV. Among the protocols tested, the protocol which presented a higher 260/280 absorbance rate, up to 4nm, and was more efficient for both human control and HPV diagnosis during amplification was the Dnaeasy.