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

Rev Soc Bras Med Trop. 2009 Nov-Dec;42(6):651-6. doi: 10.1590/s0037-86822009000600008.
[Article in Portuguese]

Abstract

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/(1/4)microl), 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/(1/4)microl). 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.

Publication types

  • English Abstract

MeSH terms

  • DNA / blood
  • DNA / isolation & purification*
  • Genotype
  • HLA Antigens / genetics*
  • Humans
  • Luminescent Measurements
  • Polymerase Chain Reaction / methods
  • Receptors, KIR / genetics*

Substances

  • HLA Antigens
  • Receptors, KIR
  • DNA