Cattle fetal sex determination using cell-free fetal DNA from maternal blood in the Mekong Delta, Vietnam

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Tran Thi Thanh Khuong
Nguyen Dang Khoa
Nguyen Van Loi
Tran Van Be Nam
Tran Gia Huy
Nguyen Trong Ngu
Lam Phuoc Thanh
Cao Thi Tai Nguyen


Cell-free fetal DNA (cffDNA) from fetal calves was isolated in the bovine maternal plasma in the fifth week of pregnancy, accounting for around 10% of the total. Recently, cffDNA has been used as a non-invasive prenatal screening technique for aneuploidy, genetic disorders in humans, early pregnancy diagnosis, and animal fetal sex determination. This study's objective was to identify the presence of cffDNA in the blood of 13 dairy cows (11 pregnant, 2 non-pregnant, and 1 unmarred) gathered from farms in Can Tho, the Mekong Delta, Vietnam. Plasma / Serum cfc-DNA Purification Midi Kit (Product # 55600) was used to separate cffDNA. Two PCR-designed primers named SRY-137 and GAPDH-109 were employed to amplify the Y-chromosome-specific sequences SRY and the housekeeping gene GAPDH, respectively. The fetal sex was indicated by PCR data corresponding to the calf's sex at birth in 11 cases. The study's findings first proved the existence of cell-free fetal DNA circulating in the plasma of a pregnant cow in Vietnam and established the calf's sex using simple, high-precision procedures. The use of cffDNA for sex screening brings up the possibility of quickly determining the gender and screening for genetic diseases in a big herd of cattle and animals with long gestation periods.

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Tran Thi Thanh Khuong, Nguyen Dang Khoa, Nguyen Van Loi, Tran Van Be Nam, Tran Gia Huy, Nguyen Trong Ngu, Lam Phuoc Thanh, & Cao Thi Tai Nguyen. (2023). Cattle fetal sex determination using cell-free fetal DNA from maternal blood in the Mekong Delta, Vietnam: Veterinary Integrative Sciences, 21(3), 657–665. Retrieved from
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Asadpour, R., Asadi, M., Jafari-Joozani, R., Hamidian, G., 2015. Ovine fetal sex determination using circulating cell-free fetal DNA (ccffDNA) and cervical mucous secretions. Asian. Pac. J. Reprod. 4(1), 65-69.

Bunnik, E.M., Kater-Kuipers, A., Galjaard, R.J. H., de Beaufort, I.D., 2020. Should pregnant women be charged for non-invasive prenatal screening? Implications for reproductive autonomy and equal access. J. Med. Ethics. 46(3), 194-198.

da Cruz, A.S., Silva, D.C., Costa, E.O.A., De M-Jr., P., da Silva, C.C., Silva, D.M., da Cruz, A.D., 2012. Cattle fetal sex determination by polymerase chain reaction using DNA isolated from maternal plasma. Anim. Reprod. Sci. 131(1-2), 49-53.

de Leon, P.M.M., Campos, V.F., Dellagostin, O.A., Deschamps, J.C., Seixas, F.K., Collares, T., 2012. Equine fetal sex determination using circulating cell-free fetal DNA (ccffDNA). Theriogenology. 77(3), 694-698.

Ershova, E., Sergeeva, V., Klimenko, M., Avetisova, K., Klimenko, P., Kostyuk, E., Veiko, N.,Veiko, R., Izevskaya, V., Kutsev, S., Kostyuk, S., 2017. Circulating cell-free DNA concentration and DNase I activity of peripheral blood plasma change in case of pregnancy with intrauterine growth restriction compared to normal pregnancy. Biomed. Rep. 7(4), 319-324.

Jimenez, D.F., Tarantal, A.F., 2003a. Quantitative analysis of male fetal DNA in maternal serum of gravid rhesus monkeys (Macaca mulatta). Pediatr. Res. 53(1), 18-23.

Jimenez, D.F., Tarantal, A.F., 2003b. Fetal gender determination in early first trimester pregnancies of rhesus monkeys (Macaca mulatta) by fluorescent PCR analysis of maternal serum. J. Med. Primatol. 32(6), 315-319.

Kadivar, A., Hassanpour, H., Mirshokraei, P., Azari, M., Gholamhosseini, K., Karami, A, 2013. Detection and quantification of cell-free fetal DNA in ovine maternal plasma; use it to predict fetal sex. Theriogenology. 79(6), 995-1000.

Kadivar, A., Rashidzadeh, H., Davoodian, N., Nazari, H., Dehghani Tafti, R., Heidari Khoei, H., Seidi Samani, H., Modaresi, J., Ahmadi, E, 2021. Evaluation of the efficiency of TaqMan duplex real‐time PCR assay for non‐invasive pre‐natal assessment of foetal sex in equine. Reprod. Domest. Anim. 56(2), 287-291.

Kazachkova, N., Gontar, J., Verlinsky, O., Ilyin, I., 2019. Successful early fetal sex determination using cell-free fetal DNA isolated from maternal capillary blood: a pilot study. Eur. J.Obstet. Gynecol. Reprod. Biol. X. 3, 100038.

Kelly, S.E., Farrimond, H.R., 2012. Non-invasive prenatal genetic testing: a study of public attitudes. Public. Health. Genom. 15(2), 73-81.

Kjeldsen-Kragh, J., Hellberg, Å, 2022. Noninvasive prenatal testing in immunohematology -clinical, technical and ethical considerations. J. Clin. Med. 11(10), 2877.

Koopman, P., Gubbay, J., Vivian, N., Goodfellow, P., Lovell-Badge, R., 1991. Male development of chromosomally female mice transgenic for Sry. Nature. 351(6322),117-121.

Lemos, D.C., Takeuchi, P.L., Rios, Á.F.L., Araújo, A., Lemos, H.C., Ramos, E.S., 2011.Bovine fetal DNA in the maternal circulation: Applications and implications. Placenta.32(11), 912-913.

Lo, Y.M.D., Hjelm, N.M., Fidler, C., Sargent, I.L., Murphy, M.F., Chamberlain, P.F., Poon,P.M.K., Redman, C.W.G., Wainscoat, J.S, 1998. Prenatal diagnosis of fetal RhD status by molecular analysis of maternal plasma. N. Engl. J. Med. 339(24),1734-1738.

Thuan, N.K., Lam, N.T., Chien, N.T.P., Khanh, N.P., Bich, T.N., 2022. Prevalence of antibiotic resistance genes and genetic relationship of Escherichia coli serotype O45, O113,O121, and O157 isolated from cattle in the Mekong Delta, Vietnam. Vet. Integr. Sci. 20(3), 695-707.

Ristanic, M., Stanisic, L., Maletic, M., Glavinic, U., Draskovic, V., Aleksic, N., Stanimirovic, Z., 2018. Bovine foetal sex determination - Different DNA extraction and amplification approaches for efficient livestock production. Reprod. Domest. Anim. 53(4),947-954.

Robinson, T.L., Sutherland, I.A., Sutherland, J., 2007. Validation of candidate bovine reference genes for use with real-time PCR. Vet. Immunol. Immunopathol. 115(1-2), 160-165.

Stévant, I., Papaioannou, M.D., Nef, S., 2018. A brief history of sex determination. Mol. Cell.Endocrinol. 468, 3-10.

Stoops, M.A., Winget, G.D., De Chant, C.J., Ball, R.L., Roth, T.L, 2018. Early fetal sexing in the rhinoceros by detection of male-specific genes in maternal serum. Mol. Reprod. Dev. 85(3), 197-204.

Tonekaboni, F.R., Narenjisani, R., Staji, H., Ahmadi-Hamedani, M., 2020. Comparison of cell-free fetal DNA plasma content used to sex determination between three trimesters of pregnancy in torkaman pregnant mare. J. Equine Vet. Sci. 95, 103273.

Vincze, B., Gáspárdy, A., Biácsi, A., Papp, E.Á., Garamvölgyi, L., Sós, E., Cseh, S., Kovács,G., Pádár, Z., Zenke, P., 2019. Sex determination using circulating cell-free fetal DNA in small volume of maternal plasma in elephants. Sci. Rep. 9(1), 15254.

Wang, G., Cui, Q., Cheng, K., Zhang, X., Xing, G., Wu, S., 2010. Prediction of fetal sex by amplification of fetal dna present in cow plasma. J. Reprod. Dev. 56(6), 639-642.

Yasmin, L., Takano, J.I., Nagai, Y., Otsuki, J., Sankai, T., 2015. Detection and quantification of male-specific fetal DNA in the serum of pregnant cynomolgus monkeys (Macaca fascicularis). Comp Med. 65(1), 70-76.