Interpretation of the results of quantitative genetic analysis based on the approximation of the kinetic curve of the polymerase chain reaction in real time

A quantitative polymerase chain reaction in real time (real time PCR) has been implemented on polymer microfluidic devices made of polycarbonate and polypropylene. The threshold cycle is determined based on the inflection point of the first-order logistic growth function, which reliably approximates the PCR curve in the absence of interfering factors, for example, the presence of bubbles in the reaction chamber. Use of statistical criteria (generalized Student's criterion, one-way analysis of variance) revealed the insignificance of the influence of the polymer type on the estimation of the position of the threshold cycle in terms of the previously selected algorithm for its search. When using an alternative algorithm for finding the threshold cycle based on the plotting of a tangent to the kinetic curve, in some cases there is a significant influence of the polymer type on the estimation of the position of the threshold cycle and, as a consequence, on the result of quantitative analysis. Proposed and discussed are algorithms for detecting bubbles in the reaction chamber based on the detection of a discrepancy in the measurement sequence associated with both estimates of the approximating dependence's parameters and the characteristics of the time series formed by approximation errors.