A novel quantitative real-time PCR method for the detection of mammalian and poultry species based on a shared single-copy nuclear DNA sequence.

ABSTRACT

Accurate quantification of species fractions is critical to determine meat adulteration. This study aimed to develop a novel quantitative real-time PCR (qRT-PCR) method for detection of mammalian and poultry DNA. A shared single-copy nuclear DNA sequence derived from the first exon of the LcoR gene was identified as a multi-species universal reference for a qRT-PCR assay. The conservation and copy number of the LcoR gene were evaluated among different species. The limit of detection was 0.01 ng DNA or 0.01% meat ingredient, and the limit of quantification was 0.01 ng DNA or 0.05% meat ingredient. Both the relative error (R.E.) and relative standard deviation (R.S.D.) were ≤ 25%. Moreover, modified coefficient k was introduced into this quantitative system to improve the accuracy and reliability of results, with maximum R.E. improved from 19.43% to 16.16%. The quantitative method would contribute to fighting against meat adulteration and maintaining a fair market.