Simple salting-out method for DNA extraction from formalin-fixed, paraffin-embedded tissues.
The aim of this study was to standardize a method of DNA extraction from formalin-fixed and paraffin-embedded tissues (PETs) using a salt solution to precipitate protein and isopropanol to precipitate DNA. The samples were submitted to a DNA extraction method in which two different concentrations of ammonium acetate (2 and 4M) were compared with a phenol-chloroform extraction method and with a commercial DNA isolation kit. DNA was qualified and quantified by spectrophotometer analysis, electrophoresis, and amplification by PCR. The 167 and 268bp fragments of APC and beta-globin genes, respectively, were amplified equally from DNA extracted by all tested methods and in all cases. However, the 536bp fragment of beta-globin gene was not amplified in all cases. According to our results, the extraction method using ammonium acetate proved to be simple and suitable for obtaining DNA of good quality, which can be easily amplified by PCR.
Rivero
Elena R C
ER
Department of Pathology, Health Science Center, Federal University of Santa Catarina, Florianópolis, SC, Brazil.
Neves
Adriana C
AC
Silva-Valenzuela
Maria G
MG
Sousa
Suzana O M
SO
Nunes
Fabio D
FD
D000085:Acetates / Q000737:chemistry; D002294:Carcinoma, Squamous Cell; D002725:Chloroform / Q000737:chemistry; D004273:DNA, Neoplasm / Q000302:isolation & purification*; D003902:Detergents / Q000737:chemistry; D005404:Fixatives; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D006965:Hyperplasia; D009062:Mouth Neoplasms / Q000737:chemistry; D016612:Paraffin Embedding; D010636:Phenols / Q000737:chemistry; D016133:Polymerase Chain Reaction / Q000379:methods; D016707:Tissue Fixation / Q000379:methods*
D000085:Acetates; D004273:DNA, Neoplasm; D003902:Detergents; D005404:Fixatives; D010636:Phenols; D005557:Formaldehyde; D002725:Chloroform; C018824:ammonium acetate