The DNA isolation method has effect on allele drop-out and on the results of fluorescent PCR and DNA fragment analysis.
BACKGROUND
The quality and the quantity of isolated DNA have an effect on PCR amplifications.


METHODS
The authors studied three DNA isolation protocols (resin binding method using fresh and frozen amniotic fluid samples, and silica adsorption method using fresh samples) on the quantity and on the quality of the isolated DNA. Amniotic fluid samples were obtained from 20 pregnant women. The isolated DNA concentrations were determined by real-time fluorimeter using SYBRGreen I method. Each sample was studied for the presence of 8 STR markers. The authors compared the number of the detected alleles, electrophoretograms and peak areas.


RESULTS
There was a significant difference between the concentration of the obtained DNA and in the peak areas between the three isolation protocols. The numbers of detected alleles were different, we observed the most allele drop outs in the resin type DNA isolation protocol from the fresh sample (detected allele numbers 182), followed by resin binding protocol from the frozen samples (detected allele number 243) and by the silica adsorption method (detected allele number 264).


CONCLUSIONS
The authors demonstrated that the DNA isolation method has an effect on the quantity and quality of the isolated DNA, and on further PCR amplifications.
Genetic Laboratory, 1st Dept. of Obstetrics and Gynecology, Semmelweis University, Budapest, Hungary. nabal@noi1.sote.hu
Bálint
B
Nagy
Zoltán
Z
Bán
Zoltán
Z
Papp
D000327:Adsorption; D000328:Adult; D000483:Alleles; D000653:Amniotic Fluid; D004247:DNA / Q000032:analysis / Q000302:isolation & purification*; D005260:Female; D005456:Fluorescent Dyes; D006801:Humans; D008722:Methods; D016133:Polymerase Chain Reaction / Q000592:standards*; D011247:Pregnancy; D012117:Resins, Synthetic; D012822:Silicon Dioxide
D005456:Fluorescent Dyes; D012117:Resins, Synthetic; D012822:Silicon Dioxide; D004247:DNA