PCR-based sexing of caprine embryos using Sry/Aml-X duplex PCR and Sry/DVEPC041 duplex PCR

RAMLI BIN ABDULLAH; CHOON YEN ALICE PHUA; MOHAMMAD MIJANUR RAHMAN; WAN EMBONG WAN KHADIJAH

doi:10.56093/ijans.v87i8.73421

Authors

RAMLI BIN ABDULLAH University of Malaya, Kuala Lumpur 50603, Malaysia
CHOON YEN ALICE PHUA University of Malaya, Kuala Lumpur 50603, Malaysia
MOHAMMAD MIJANUR RAHMAN University of Malaya, Kuala Lumpur 50603, Malaysia
WAN EMBONG WAN KHADIJAH University of Malaya, Kuala Lumpur 50603, Malaysia

DOI:

https://doi.org/10.56093/ijans.v87i8.73421

Keywords:

Caprine, Polymerase chain reaction, Sex determination

Abstract

This study presents the development of a PCR-based method for sex determination of caprine embryos by Sry and Aml-X gene detection. DNA sequencing of the PCR products amplified from the target loci confirmed that these amplicons were indeed amplified from portions of the caprine Sry and Aml-X genes. Furthermore, the primer sets were found to be specific for sexing of caprine of various breeds. An alternative PCR-sexing protocol was also developed to verify embryo sexing results and it was a pit-stop using Sry and DVEPC041 primer sets. For the pitstop PCR protocol, the Sry primer was best added after the first 6 cycles to obtain bands of equal intensity at 116 bp and 160 bp when identifying the male sex. However, application of the same pit-stop PCR protocol using genomic DNA of various caprine breeds also produced acceptable and accurate sex identification despite showing slight diversion of the expected result.

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References

Bredbacka P, Kankaanpaa A and Peippo J. 1995. PCR-sexing of bovine embryos: a simplified protocol. Theriogenology 44: 167–76. DOI: https://doi.org/10.1016/0093-691X(95)00166-6

Chen A Q, Xu Z R and Yu S D. 2007. Sexing goat embryos by PCR amplification of X- and Y- chromosome specific sequence of the amelogenin gene. Asian Australasian Journal of Animal Sciences 20: 1689–93. DOI: https://doi.org/10.5713/ajas.2007.1689

Dervishi E, Sanchez P, Alabart J I, Cocero M J, Folch J and Calvo J H. 2011. A suitable duplex PCR for ovine embryo sex and genotype of PrnP gene determination for MOET-based selection programmes. Reproduction in Domestic Animals 46: 999–1003. DOI: https://doi.org/10.1111/j.1439-0531.2011.01774.x

Hasler J F, Cardey E, Stokes J E and Bredbacka P. 2002. Nonelectrophoretic PCR-sexing of bovine embryos in a commercial environment. Theriogenology 58: 1457–69. DOI: https://doi.org/10.1016/S0093-691X(02)01044-0

Lopes R F F, Senna J P M, Chies J M and Rodrigues J L. 1999. Pit-stop PCR: an approach to increase initial product yield of multiplex PCR. Biotechniques 26: 638–39. DOI: https://doi.org/10.2144/99264bm09

Nor Farizah A H, Rahman M M, Wan Khadijah W E and Abdullah R B. 2015. Effect of repeated laparoscopic ovum pick-up on yield and quality of oocytes in goats. Malaysian Journal of Animal Science 18: 55–59.

Phua A C Y, Rahman M M, Wan Khadijah W E and Abdullah R B. 2015. Influence of in vitro maturation duration on embryo yield in goats. Malaysian Journal of Animal Science 18: 23– 28.

Phua A C Y, Abdullah R B and Mohammed Z. 2003. A PCR- based sex determination method for possible application in caprine gender selection by simultaneous amplification of the Sry and Aml-X genes. Journal of Reproduction and Development 49: 307–11. DOI: https://doi.org/10.1262/jrd.49.307

Saberivand A and Ahsan S. 2016. Sex determination of ovine embryos by SRY and amelogenin (AMEL) genes using maternal circulating cell free DNA. Animal Reproduction Science 164: 9–13. DOI: https://doi.org/10.1016/j.anireprosci.2015.10.011