Determination of content of camel milk in adulterated milk samples by normalized real-time polymerase chain reaction system based on single-copy nuclear genes.

Abstract

Compared with the traditional qualitative polymerase chain reaction (PCR), which only identifies the category of species, the quantitative PCR method provides a value, which is very important for appropriate penalty enforcement according to the extent of adulteration. However, most of the current quantitative PCR methods are based on mitochondrial genes, expressing different copy numbers in different cells and reducing the accuracy of quantitative results. In this study, single-copy nuclear housekeeping genes, instead of multicopy mitochondrial genes, were selected as both camel species-specific and reference genes to develop a novel normalized PCR system. This system had an excellent linear correlation (R2 = 0.9614) between camel milk content and Ct ratio (specific/reference genes), and allowed quantitative determination of the content of camel milk in adulterated milk samples. The accuracy was effectively validated using simulated adulterated samples with recoveries ranging from 90% to 120% and coefficient of variation less than 10%, exhibiting sufficient parameters of trueness and precision. The normalized PCR system based on single-copy nuclear genes is a simple, rapid and reliable method for the determination of the content of camel milk in adulterated milk samples, and also provides technical support for appropriate penalty enforcement. © 2020 Society of Chemical Industry.