DEVELOPMENT OF OOCYTE MATURATION MEDIUM FOR BOVINE ASSISTED REPRODUCTIVE TECHNOLOGY
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Date
2022
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ICAR-NDRI, KARNAL
Abstract
Oocyte maturation is an inevitable part of assisted reproductive technology (ART) and indigenously developed medium for in vitro maturation is a need of the hour. For the development of oocyte maturation medium for bovine ART, this study was carried out with the following objectives: i) To optimize the culture medium for in vitro maturation of buffalo oocytes, 2) To examine the development competence of in vitro produced embryos using an optimized maturation medium and 3) To study the effect of optimized maturation medium on pluripotency, development and apoptosis-related genes in produced embryos. For this oocytes were cultured in four different maturation media, of which two were formulated in our lab (L-OMM-1 and L-OMM-2) and two commercially available media (C-OMM-1 and C-OMM-2). The buffalo oocytes were cultured for 24 h in CO2 incubator at 5% CO2 in air at 38.5°C for in vitro maturation. The polar body extrusion showed that L-OMM-1 had 79.52±1.48% maturation rate which was significantly higher (P<0.05) than L-OMM-2 (31.51±0.6%); and at par with the maturation rate of C-OMM-1 (82.57±1.96%), but lower (P<0.05) than C-OMM-2 (87.22±1.91%). The oocytes in L-OMM-1 (n=306) and C-OMM-1 (n=288) media were matured in vitro and subjected to IVF and IVC in similar media for both the groups. The cleavage rate observed between above two groups had no difference (64.96 ± 1.33 vs 64.40 ± 2.12) whereas blastocyst rate was found to be at par in both the media (P>0.05) (16.31 ± 1.41 vs 17.71 ± 0.24). TUNEL assay showed no significant (P>0.05) difference in apoptotic index of both the blastocyst groups produced in L-OMM-1 (4.95±0.17%) and C-OMM-1 (4.41±0.19%). The total cell number in blastocyst was higher in L-OMM-1 media (191.88±17.05) compared to C-OMM-1 media (171.63±12.48) but differ non-significantly (P>0.05). The gene expression study revealed that SOD2, BCL-XL, BMP15 and HPRT genes were significantly up-regulated (P<0.05), P53, BAX, GLUT1 and OCT4 were significantly down-regulated (P<0.05), whereas no change in expression of BAD, IGF2, C-MYC in blastocyst obtained from L-OMM-1 compared to C-OMM-1. This all suggests that the lab-formulated medium has competence for in vitro oocyte maturation and subsequent to embryo development as compared to commercial media. In conclusion, the lab-formulated oocyte maturation media (L-OMM-1) can be used for in vitro maturation of oocytes.