Avaliação de dois métodos de extração de DNA de material parafinado para amplificação em PCR

Abstract

BACKGROUND: There are several methods for DNA extraction from formalin-fixed paraffin-embedded tissues reported in the literature. High success rate on this procedure is important for the use of molecular diagnostic methods based on the polymerase chain reaction (PCR). OBJECTIVE: Two methods of DNA extraction from paraffin-embedded samples were tested aiming at PCR amplification of genomic DNA. MATERIAL AND METHOD: Thirty-five samples were obtained from patients with squamous cell carcinoma of mouth floor treated at the Oral Oncology Center in Universidade Estadual Paulista. The DNA extraction methods included: 1. proteinase K digestion followed by Chelex 100® (BioRad) purification and 2. QIAamp DNA minikit® system (Qiagen). Purified DNA was quantified by spectrophometry and a beta-globin gene fragment was amplified using PCR. RESULTS: The DNA concentration from samples applied to the first method presented an average of 120.62 ng/µl and absorbance ratio 260/280 varying between 0.8 and 1.41. From the samples extracted using the second procedure, the mean DNA concentration was 67.38 ng/µl, with absorbance ratio varying between 1.11 and 2.53. DNA samples were submitted to PCR and from 35 samples extracted with both methods, respectively, 29 and 30 were successfully amplified for the beta-globin gene. CONCLUSION: Both methods used to obtain genomic DNA from formalin-fixed paraffin-embedded tissues presented similar performance, revealing their potential to be included in diagnosis of molecular biology, as well as in retrospective studies using archived paraffin-embedded samples.