Augmentation of Temperatures, Media and Cryoprotectants on in vitro Fertilization of Pig Oocytes
- 1 Agricultural Research Council, Animal Production, Germplasm Conservation and Reproductive Biotechnologies, Private Bag X 2, Irene, South Africa
- 2 Department of Animal Sciences, Faculty of Science, Tshwane University of Technology, Private Bag X 680, Pretoria, South Africa
Abstract
There are numerous challenges to the large-scale production of pig embryos using in vitro procedures. The aim of the present study was to compare the different holding temperatures, in vitro maturation media, and permeating cryoprotectants in the in vitro fertilization of pig oocytes. Ovaries were obtained from the local slaughterhouse, oocytes were retrieved by means of aspiration and slicing techniques and categorized into grades A, B, and C. Good quality oocytes were stored in an Eppendorf tube containing a holding solution at 5, 18, 24, and 38.5°C for 5, 30, 60, and 120 min, respectively. Oocytes were matured for 44 h and then checked for polar body extrusion. Matured oocytes were exposed to cryoprotectant Toxicity Test (TT1) with 7.5% of DMSO + 7.5% of EG and (TT2) with 15% of DMSO + 15% EG. Oocytes were then in vitro fertilized with fresh semen, incubated for 24 h and the pronucleus was checked. The GLM procedure was analyzed using Analysis of Variance (ANOVA) and treatment means were compared with the Least Significance Difference (LSD) test. The results revealed that at 38.5°C, the Cumulus Oocytes Complexes (COCs) expansion rate was significantly (p<0.05) higher (75.8±14.0), no damaged cytoplasm and the polar body was higher (25.5±7.6) compared to all other treatments. Oocytes polar body extrusion were (32.8±13.6, 25.8±9.1 and 11.5±6.9) for NCSU-23, NCSU-37 and TCM-199 respectively. There was a significant (p<0.05) difference in the total fertilization rate of the control (75.8±17.2) and the combination of DMSO and EG (50.3±21.4). Pig oocytes are very sensitive to lower temperatures after IVM. The NCSU-23 and NCSU-37 media enhance pig oocytes polar body extrusion. The combination of DMSO and EG affected the pronucleus since the cryoprotectants have toxicity when used on pig oocytes after maturation.
DOI: https://doi.org/10.3844/ajavsp.2024.247.256
Copyright: © 2024 Tumelo Thelma Maduwa, Masindi Lottus Mphaphathi and Tshimangadzo Lucky Nedambale. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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Keywords
- IVF
- Oocytes
- Permeating Cryoprotectants
- Polar Body
- Pronuclear