Evaluation of the efficiency of TaqMan duplex real-time PCR assay for non-invasive pre-natal assessment of foetal sex in equine

Accurate diagnosis of foetal sex in pregnant mare is helpful for many breeders, both for private or commercial purposes. In this study, in order to pre-natal foetal sexing in equine, we used TaqMan duplex real-time PCR to detect the specific regions of SRY and TSPY genes on extracted cell-free foetal DNA from maternal blood. Peripheral blood samples from 50 pregnant Arabian mares with singleton foetuses were collected. Cell-free foetal DNA was extracted from maternal plasma, and duplex real-time PCR assays were performed with TaqMan probes and primers. Amplification of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was used as control of DNA extraction procedure. From the 50 sampled mares, 28 cases had female and 22 mares had male foetuses. The final results for 46 samples were conclusive, and from them, 43 cases were predicted correctly. Sensitivity, specificity and accuracy of the test were 90.48%, 96% and 93.48%, respectively. In conclusion, a TaqMan duplex real-time PCR was set up to pre-natal detection of foetal sex in equine. The method was fast and decreased the false-positive and false-negative results. The technique can be used as a routine procedure in farms by collecting only a blood sample.