화학공학소재연구정보센터
Biochemical and Biophysical Research Communications, Vol.299, No.5, 715-722, 2002
Kinetic characterisation of primer mismatches in allele-specific PCR: a quantitative assessment
A novel method of estimating the kinetic parameters of Taq DNA polymerase during rapid cycle PCR is presented. A model was constructed using a simplified sigmoid function to represent substrate accumulation during PCR in combination with the general equation describing high substrate inhibition for Michaelis-Menten enzymes. The PCR progress curve was viewed as a series of independent reactions where initial rates were accurately measured for each cycle. Kinetic parameters were obtained for allele-specific PCR (AS-PCR) amplification to examine the effect of mismatches on amplification. A high degree of correlation was obtained providing evidence of substrate inhibition as a major cause of the plateau phase that occurs in the later cycles of PCR. (C) 2002 Elsevier Science (USA). All rights reserved.