Identifying freemartins using qualitative and quantitative PCR of the SRY gene in cattle and water buffalo species

Abstract Freemartin is a disorder where physical and hormonal characteristics from a male fetus are transferred to a female fetus through the placental anastomosis. This causes the female twin to be mostly infertile, leading farmers to cull them due to the lack of a reliable diagnostic approach, causing economic and genetic losses. The study aims to develop an accurate diagnostic tool to identify freemartinism using comparative assessment of the SRY gene through qualitative (PCR) and quantitative (qPCR) methods and relate to their consequent fertility status. Blood samples from suspected freemartins were collected, and DNA and RNA were extracted and analyzed to verify freemartinism using the SRY gene. The results show that PCR and qPCR techniques can identify freemartinism cases using the SRY gene. Using PCR, the SRY gene was absent in fertile freemartin, and non-fertile freemartin and freemartin singleton were positive with SRY. qPCR showed that the relative content of SRY was 4.84–48% more in freemartins than in co-twin males and undetected in fertile freemartins. The small yet detectable amount of the SRY gene in the freemartin samples explains possible chimerism, contamination of male hormones, and underdevelopment of gonads and reproductive tracts, which could cause infertility in animals. Therefore, this developed method for detecting freemartinism can help farmers make informed decisions regarding breeding practices to optimize fertility and productivity in their herds.