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Effect of Fresh and Frozen Semen on in Vitro Fertilization and Subsequent Development of Goat Embryos

S. Saha
Department of Biotechnology, Bangladesh agricultural university, Mymensingh, Bangladesh.

M.A.M.Y. Khandoker
Department of Animal Breeding and Genetics, Bangladesh agricultural university, Mymensingh, Bangladesh.

L.Y. Asad
Department of Animal Breeding and Genetics, Bangladesh agricultural university, Mymensingh, Bangladesh, Department of Animal Nutrition, Genetics and Breeding, Sher? e - Bangla Agricultural University, Dhaka, Bangladesh.

A.M.M.T. Reza
Department of Animal Breeding and Genetics, Bangladesh agricultural university, Mymensingh, Bangladesh.

A. Hoque
Department of Animal Breeding and Genetics, Bangladesh agricultural university, Mymensingh, Bangladesh.

Abstract/ Executive Summary:

This study was undertaken to compare the efficiency of fresh and frozen-thawed buck semen on in vitro fertilization (IVF) of goat oocytes. Cumulus oocytes comp lexes (COCs) were collect ed by aspiration of 2-6 mm diameter follicles, which were obtained from slaughterhouse. Upon grading, only normal quality COCs were maturated in TCM-199 for 48 hours. The percentage of COCs reached to the M-II stage was 61.41 ± 1.97%. The matured COCs were fertilized for 5 hours in Brackett and Oliphant media using fresh and fro- zen thawed semen separately. After fertilization the oocytes were cultured in TCM-199 for 48 hours to ob- serve the cleavage rate. The maturation, fertilization and culture were performed in an incubator at 38.5 ⁰C with 5% CO₂ in humidified air. After fertilization cleavage rates were observed to check the fitness of zygotes to be morula and blastocyst. It was observed that the rates of normal fertilization (2 PN formations) for fresh and frozen semen were 36.02 ± 2.79 and 34.73 ± 2.58%, respectively and the cleavage rates were 25.19 ± 2.5 and 21.01 ± 2.8%, respectively. No significant differences (P>0.05) was observed between fresh and frozen semen in the efficiency of in vitro fertilization and subsequent development of goat embryos. It can be concluded that, both fresh and frozen semen can be used for IVF and subsequent development of goat embryos.

Keywords: Goat, Frozen semen, In vitro fertilization, Goat oocytes

Location: Bangladesh agricultural university, Mymensingh.

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Goat

Objectives:

To compare the efficiency of fresh and frozen buck semen on in vitro fertilization (IVF) of goat oocytes.

Methodology:

The experiment was conducted at the reproductive biotechnology laboratory, department of animal breeding and genetics, Bangladesh agricultural university, Mymensingh. A total of 195 ovaries from goats of unknown reproductive history were collected from local slaughterhouse. The ovaries were kept in collection vial containing 0.9% physiological saline in a thermo flask at 25 ⁰C to 30 ⁰C and trans- ported to the laboratory within 4 to 5 hours of slaughter. The ovaries were rinsed thoroughly by physiological saline solution at 25 ºC. In the laboratory, each ovary was trimmed to remove the surrounding tissues and overlying bursa. After collection and trimming ovaries were evaluated on the basis of the corpus luteum (CL) present and absent. After collection, trimming and washing of ovaries, COCs were collected by aspiration from 2-6 mm diameter follicles using 10 mL syringe with 18 gauge needle. The COCs were then classified into 4 grades as described. The grades: grade A) oocytes completely surrounded by cumulus cells; grade B) oocytes partially sur- rounded by cumulus cells; grade C) oocytes not surrounded by cumulus cells and grade D) degeneration observed both in oocytes and cumulus cells. The grade A and B were considered as normal and grade C and D as abnormal COCs. Normal quality COCs (A and B grade) were washed 2-3 times separately in D-PBS and finally into the maturation media (TCM-199) and placed in to the droplets containing TCM-199. The droplets were then kept in a CO₂ incubator at 38.5 ⁰ C with 5% carbon dioxide in humidified air for 48 hours. After 48 hours of maturation, the half of the matured COCs was proceed to fertilization w ith fresh semen and remaining half of the matured COCs was proceed to fertilization with frozen semen in Brackett and Oliphant (BO) medium in CO₂ incubator at 38.5 ⁰C with 5% carbon dioxide in humidified air for 6 hours. Then the pronuclei developments were observed separately for fresh and frozen semen. After 5 hours incubation, the fertilized ova were taken from the semen drops with cumulus cells by using the appropriate micropipette. Then, they we rewashed three times in pre-incubated medium (TCM-199) and were transferred to other culture drop (100µL) of TCM-199. The dish was then kept in the CO₂ incubator at 38.5 ^oC with 5% CO₂ in air. The development rates were checked after 48 hours of culture. The cleavage rates were recorded after 48 hours of culture. The data generated from this study were entered in micro- soft excel worksheet, organized and processed for analysis. Analysis was performed with the help of statistical analysis system.

Source of Information: Iranian Journal of Applied Animal Science (2014) 4(2), 325-330

Article Link (Online Journal):

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Considering the effect of semen on in vitro fertilization and culture of goat oocytes, it can be concluded that both fresh and frozen semen can be used on in vitro fertilization of goat oocytes and subsequent development of goat embryos.

Knowledge Management: Journal