DNA extraction from coagulated human blood for application in genotyping techniques for human leukocyte antigen and immunoglobulin-like receptors

Revista da Sociedade Brasileira de Medicina Tropical

Braga, Marco Antônio¹  Guelsin, Gláucia Andréia¹  Melo, Fabiano Cavalcante de¹  Moliterno, Ricardo Alberto¹  Visentainer, Jeane Eliete Laguila¹  Cardozo, Daniela Maira¹  Secretaria de Saúde do Estado do Paraná, Maringá¹  Souza, Cleonice de¹  Clementino, Samaia Laface¹  Universidade Estadual de Maringá, Maringá¹ 

关键词: DNA extraction;    Coagulated blood;    Molecular biology;    Standardization;   

DOI  :  10.1590/S0037-86822009000600008

学科分类：农业科学（综合）

来源: Sociedade Brasileira de Medicina Tropical

【 摘 要 】

The objective of this study was to standardize a method for extracting high-quality DNA from samples of coagulated blood. Forty-eight samples of human coagulated blood were used for DNA extraction by means of the EZ-DNA® commercial kit (Biological Industries, Beit Haemek, Israel), the Neoscience® column kit (One Lambda Inc., San Diego, CA, USA) and a modified salting-out method. Only the salting-out method was able to extract high concentrations of DNA (mean, 180 ng/¼l), which were measured using the Qubit® fluorescence detector (Invitrogen, USA). This method enabled amplification of HLA (human leukocyte antigen) genes using the Luminex PCR-SSO (polymerase chain reaction - sequence-specific oligonucleotide) technology, which demands good quality DNA, and amplification of KIR (killer-cell immunoglobulin-like receptor) genes using an in-house PCR-SSP (polymerase chain reaction - sequence-specific primer) technique, which demands a specific concentration of DNA (10 ng/¼l). We concluded that the modified salting-out technique was very efficient, simple and fast for DNA extraction from human coagulated blood samples, with the aim of genotyping the HLA and KIR genes.

【 授权许可】

Unknown