Title: Comparison of DNA Isolation Kits to Extract DNA from Whole Blood Samples
Volume: 7
Issue: 2
Author(s): Robert A.M. Op den Buijsch, Johan E. de Vries, Joop ten Kate, Petal A.H.M. Wijnen, Snjezana Rothkrantz-Kos, Marja P. van Dieijen-Visser and Otto Bekers
Affiliation:
Keywords:
Cycle theshold (Ct), DNA isolation kit comparison, UV spectrophotometry, real-time PCR, White blood cells (WBC)
Abstract: Genomic DNA from WBCs is widely often used for PCR. Although kits for DNA isolation are in common use, there is scarce information about their performance and about the PCR quality of the genomic DNA obtained. Hence, three different kits, QIAamp blood mini kit, High Pure PCR Template Preparation Kit and the Puregene DNA isolation kit were evaluated on these aspects. Genomic DNA was isolated from whole blood samples with WBC counts ranging from 0.5 to 20*109 WBC/L. The WBC count was used to calculate the amount of genomic DNA. The actual amount of genomic DNA isolated, was determined spectrophotometrically. The DNA extraction efficiency was obtained by dividing the actual amount of DNA by the calculated amount yielded. PCR quality was analysed by measuring Cycle threshold (Ct) values with a quantitative real-time PCR β-globin assay. The extraction efficiency of the three DNA isolation kits was 20% to 40%. Spectrophotometrically determined DNA concentrations correlated inversely with Ct values, regardless of the DNA isolation kit applied, whereas the strongest correlation was obtained for the Puregene DNA isolation kit. WBC counts also correlated inversely with Ct values and here the strongest correlation was found for the QIAamp blood mini kit. In conclusion, the overall performance of the DNA isolation kits was quite comparable (DNA recoveries of 20-40%), albeit the QIAamp blood mini kit yielded the most reproducible extraction efficiencies and lowest Ct values within every WBC count category.
