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Research Article

volume 54 issue 10 (october 2020) : 1210-1213,   Doi: 10.18805/ijar.B-3886

In vitro Development of Preimplantation Caprine Embryo using Cryopreserved Black Bengal Buck Semen

R
Rohit Kumar
P
P. Chandra
P
P. Konyak
M
M. Karunakaran
A
A. Santra
S
Subrata K. Das
1Animal Biotechnology Lab, ICAR-National Dairy Research Institute, Eastern Regional Station, Kalyani-741 235, West Bengal, India.
Cite article:- Kumar Rohit, Chandra P., Konyak P., Karunakaran M., Santra A., Das K. Subrata (2020). In vitro Development of Preimplantation Caprine Embryo using Cryopreserved Black Bengal Buck Semen. Indian Journal of Animal Research. 54(10): 1210-1213. doi: 10.18805/ijar.B-3886.

ABSTRACT

The aim of the present study was to produce goat embryo in different culture media through in vitro fertilization using cryopreserved black Bengal buck semen. Total 1265 fresh cumulus oocyte complexes (COCs) were collected by aspiration method with a 19 gauge hypodermic needle, washed 5-6 times and cultured in maturation media maintaining 5% CO2 level at 38.5ºC with maximum humidity in a incubator. After 27 h of incubation cumulus cells were stripped off from matured oocytes and transferred to acidified Tyrode’s medium for zona thinning and co-incubated with in vitro capacitated sperms for fertilization in Fert-BO media. In the experiment I, fresh buck semen and in experiment II, frozen buck semen was used for in vitro fertilization after in vitro processing. After 5 h of co-incubation, presumptive zygotes were washed and co-incubated with oviductal cells in three different culture media (RVCL, mSOF, KSOM) for further development. In fresh group cleavage (%) were 37.76 ± 2.98, 39.60 ± 1.75, 29.01 ± 1.74, and morula formation (%) were 7.72 ± 3.38, 6.03 ± 1.29, and 3.00 ± 3.00 in RVCL, mSOF and KSOM media respectively. However, in frozen group cleavage (%) were 29.17 ± 2.56, 27.70 ± 2.31, 24.17 ± 1.44 in RVCL, mSOF and KSOM media respectively and morula formation (%) was 2.93 ± 0.97 only in RVCL media. These results indicate that cryopreserved black Bengal buck semen have competence to produce embryos and could be used for embryo development in RVCL media through in vitro fertilization. 

REFERENCES

  1. Bracket, B.G. and Oliphant, G. (1975). Capacitation of rabbit spermatozoa in vitro. Biology of Reproduction. 12: 260-274.
  2. Danilda. (2000). Technical aspect of in vitro embryo production. Duran Embryo Biotechnology Laboratory, Philippine Carabao Center, National Headquarters abd gene pool Munoz, Nueva Eceija 3120 Philipine.
  3. Edwards, R.G. and Beard, H.K. (1999). Is the success of human IVF more a matter of genetics and evolution than growing blastocysts? Human Reproduction. 14: 1-4.
  4. Freitas, J.F.V. and Melo, M.L. (2010). In vitro embryo production in small ruminants. Revista Brasileira de Zootecnia. 39: 409-413.
  5. Hughes, C.M., Lewis, S.E., McKelvey-Martin, V.J., Thompson, W. A. (1996). A comparison of baseline and induced DNA damage in human spermatozoa from fertile and infertile men, using a modified comet assay. Molecular Human Reproduction. 2: 613-619.
  6. Izquierdo, D., Villamediana, P., Paramio, M.T. (1999). Effect of culture media on embryo development from prepubertal goat IVM–IVF oocytes, Theriogenology. 52:847-861.
  7. Jena, M.K., Malakar, D., De, A.K., Garg, S., Akshey, Y.S., Dutta, R., Sahu, S., Mohanty, A.K., Kaushik, J.K. (2012). Handmade cloned and parthenogenetic goat embryos-a comparison of different culture media and donor cells. Small Ruminant Research. 105:255-262.
  8. Kharche, S.D., Yadav, E.N., Goel, A.K., Jindal, S.K., Sinha, N.K. (2008a). Influence of culture media on in-vitro fertilization of goat oocytes. Indian Journal of Animal Sciences. 78: 1075-1077.
  9. Kharche, S.D., Goel, A.K., Jindal, S.K., Yadav, P., Sinha, R. and Sinha, N.K. (2009). Effect of serum albumin supplementation on in- vitro capacitaion and fertilization of caprine oocytes. Small Ruminant Research. 81: 85-89.
  10. Konyak, P., Mandal, A., Mondal, M., Bhakat, C., Das, S.K., Rai, S., Ghosh, M.K. and Karunakaran, M. (2018). Preservation of black Bengal buck semen in soybean lecithin based chemically defined extender. Indian Journal of Animal Research. 52(8): 1151-1154.
  11. Lv, L., Yue, W., Liu, W., Ren, Y., Li, F., Lee, K.B., Smith, G.W. (2009). Effects of sperm pretreatments and in vitro culture systems on development of in vitro fertilized embryos derived from prepubertal goat oocyte in China. Asian-Australasian Journal of Animal Sciences. 22: 969-967.
  12. Rohit, K., Chandra, P., Konyak, P., Karunakaran, M., Santra, A., Das, S.K. (2017). In-vitro development of caprine embryo using cryopreserved black Bengal buck semen. Reproduction, Fertility and Development. 29(1): 188-188.
  13. Ron-el, R., Nachum, H., Herman, A., Golan, A., Caspi, E., Soffer, Y. (1991). Delayed fertilization and poor embryonic development associated with impaired semen quality. Fertility and Sterility. 55: 338–344.
  14. Saha, S., Khandoker, M.A.M.Y., Asad, L.Y., Reza, A.M.M.T., Hoque, A. (2014). Effect of Fresh and Frozen Semen on in vitro Fertilization and Subsequent Development of Goat Embryos. Iranian Journal of Applied Animal Sciences. 4: 325-330.
  15. Singh, A.P., Kumar, D., Gopalakrishna R, Rakesh, R., Pandey, S.K., Sarkhel, B.C. (2016). Comparison of culture media for their effects on development of caprine IVF embryos using fresh and cryopreserved semen. Indian Journal of Animal Research. 50(6): 846-850.
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    Research Article

    volume 54 issue 10 (october 2020) : 1210-1213,   Doi: 10.18805/ijar.B-3886

    In vitro Development of Preimplantation Caprine Embryo using Cryopreserved Black Bengal Buck Semen

    R
    Rohit Kumar
    P
    P. Chandra
    P
    P. Konyak
    M
    M. Karunakaran
    A
    A. Santra
    S
    Subrata K. Das
    1Animal Biotechnology Lab, ICAR-National Dairy Research Institute, Eastern Regional Station, Kalyani-741 235, West Bengal, India.
    Cite article:- Kumar Rohit, Chandra P., Konyak P., Karunakaran M., Santra A., Das K. Subrata (2020). In vitro Development of Preimplantation Caprine Embryo using Cryopreserved Black Bengal Buck Semen. Indian Journal of Animal Research. 54(10): 1210-1213. doi: 10.18805/ijar.B-3886.

    ABSTRACT

    The aim of the present study was to produce goat embryo in different culture media through in vitro fertilization using cryopreserved black Bengal buck semen. Total 1265 fresh cumulus oocyte complexes (COCs) were collected by aspiration method with a 19 gauge hypodermic needle, washed 5-6 times and cultured in maturation media maintaining 5% CO2 level at 38.5ºC with maximum humidity in a incubator. After 27 h of incubation cumulus cells were stripped off from matured oocytes and transferred to acidified Tyrode’s medium for zona thinning and co-incubated with in vitro capacitated sperms for fertilization in Fert-BO media. In the experiment I, fresh buck semen and in experiment II, frozen buck semen was used for in vitro fertilization after in vitro processing. After 5 h of co-incubation, presumptive zygotes were washed and co-incubated with oviductal cells in three different culture media (RVCL, mSOF, KSOM) for further development. In fresh group cleavage (%) were 37.76 ± 2.98, 39.60 ± 1.75, 29.01 ± 1.74, and morula formation (%) were 7.72 ± 3.38, 6.03 ± 1.29, and 3.00 ± 3.00 in RVCL, mSOF and KSOM media respectively. However, in frozen group cleavage (%) were 29.17 ± 2.56, 27.70 ± 2.31, 24.17 ± 1.44 in RVCL, mSOF and KSOM media respectively and morula formation (%) was 2.93 ± 0.97 only in RVCL media. These results indicate that cryopreserved black Bengal buck semen have competence to produce embryos and could be used for embryo development in RVCL media through in vitro fertilization. 

    REFERENCES

    1. Bracket, B.G. and Oliphant, G. (1975). Capacitation of rabbit spermatozoa in vitro. Biology of Reproduction. 12: 260-274.
    2. Danilda. (2000). Technical aspect of in vitro embryo production. Duran Embryo Biotechnology Laboratory, Philippine Carabao Center, National Headquarters abd gene pool Munoz, Nueva Eceija 3120 Philipine.
    3. Edwards, R.G. and Beard, H.K. (1999). Is the success of human IVF more a matter of genetics and evolution than growing blastocysts? Human Reproduction. 14: 1-4.
    4. Freitas, J.F.V. and Melo, M.L. (2010). In vitro embryo production in small ruminants. Revista Brasileira de Zootecnia. 39: 409-413.
    5. Hughes, C.M., Lewis, S.E., McKelvey-Martin, V.J., Thompson, W. A. (1996). A comparison of baseline and induced DNA damage in human spermatozoa from fertile and infertile men, using a modified comet assay. Molecular Human Reproduction. 2: 613-619.
    6. Izquierdo, D., Villamediana, P., Paramio, M.T. (1999). Effect of culture media on embryo development from prepubertal goat IVM–IVF oocytes, Theriogenology. 52:847-861.
    7. Jena, M.K., Malakar, D., De, A.K., Garg, S., Akshey, Y.S., Dutta, R., Sahu, S., Mohanty, A.K., Kaushik, J.K. (2012). Handmade cloned and parthenogenetic goat embryos-a comparison of different culture media and donor cells. Small Ruminant Research. 105:255-262.
    8. Kharche, S.D., Yadav, E.N., Goel, A.K., Jindal, S.K., Sinha, N.K. (2008a). Influence of culture media on in-vitro fertilization of goat oocytes. Indian Journal of Animal Sciences. 78: 1075-1077.
    9. Kharche, S.D., Goel, A.K., Jindal, S.K., Yadav, P., Sinha, R. and Sinha, N.K. (2009). Effect of serum albumin supplementation on in- vitro capacitaion and fertilization of caprine oocytes. Small Ruminant Research. 81: 85-89.
    10. Konyak, P., Mandal, A., Mondal, M., Bhakat, C., Das, S.K., Rai, S., Ghosh, M.K. and Karunakaran, M. (2018). Preservation of black Bengal buck semen in soybean lecithin based chemically defined extender. Indian Journal of Animal Research. 52(8): 1151-1154.
    11. Lv, L., Yue, W., Liu, W., Ren, Y., Li, F., Lee, K.B., Smith, G.W. (2009). Effects of sperm pretreatments and in vitro culture systems on development of in vitro fertilized embryos derived from prepubertal goat oocyte in China. Asian-Australasian Journal of Animal Sciences. 22: 969-967.
    12. Rohit, K., Chandra, P., Konyak, P., Karunakaran, M., Santra, A., Das, S.K. (2017). In-vitro development of caprine embryo using cryopreserved black Bengal buck semen. Reproduction, Fertility and Development. 29(1): 188-188.
    13. Ron-el, R., Nachum, H., Herman, A., Golan, A., Caspi, E., Soffer, Y. (1991). Delayed fertilization and poor embryonic development associated with impaired semen quality. Fertility and Sterility. 55: 338–344.
    14. Saha, S., Khandoker, M.A.M.Y., Asad, L.Y., Reza, A.M.M.T., Hoque, A. (2014). Effect of Fresh and Frozen Semen on in vitro Fertilization and Subsequent Development of Goat Embryos. Iranian Journal of Applied Animal Sciences. 4: 325-330.
    15. Singh, A.P., Kumar, D., Gopalakrishna R, Rakesh, R., Pandey, S.K., Sarkhel, B.C. (2016). Comparison of culture media for their effects on development of caprine IVF embryos using fresh and cryopreserved semen. Indian Journal of Animal Research. 50(6): 846-850.
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