engleski

Significance of application of verified sets for DNA isolation in molecular diagnostics

Introduction: For the molecular diagnostic it is important to obtain integral DNA without potential inhibitors of polymerase chain reaction (PCR) method. Aim: To investigate the efficiency of DNA isolation and quality of DNA, obtained from whole EDTA-blood using two commercially procurable sets for DNA isolation, as well as standard phenol- chloroform method. Materials and methods: DNA was isolated using "High Pure PCR-Template Preparation Kit" (Roche, Germany, EU) - set 1 and "Biotools Bloodclean DNA Purification Kit" (Biotools, Spain, EU) - set 2, as well as phenol-chloroform DNA extraction method. Fifty EDTA-blood samples were analyzed. Results: Time required for DNA isolation using commercial sets 1 and 2 (45 minutes and 90 minutes per sample, respectively) compared to phenol-chloroform method (3 days per sample) was significantly shorter. Also, commercial sets were less toxic in comparison to standard method. Set 1 was the most reliable because obtained DNA was intact and its concentration was within precisely defined range. Application of set 2 resulted in a high content of broken, useless DNA. Besides, by using DNA samples, obtained by set 2, PCR reaction was inhibited, due to residual chemicals in 20% of cases. Conclusion: Since quality of commercially procurable sets for DNA isolation considerably differs, it is very important to examine the quality of obtained DNA, as well as hurtfulness of sets, and choose a test with the best characteristics for molecular diagnostic purpose.

DNA isolation ; molecular diagnostic

Abstract of the Scientific Syposium with International Participation "40 Years of Nuclear Medicine in Osijek", November 15 - 17, 2007. Osijek, Croatia.