No abstract

The aim of the present investigation was to study the effect of melatonin on in vitro maturation and subsequent embryo development of caprine oocytes. In experiment 1, 384 in vitro matured oocytes were selected and randomly divided in to two groups, viz. group 1 (control) wherein oocytes matured in maturation media without supplementation for 27 h in humidified atmosphere at 38.5°C with 5% CO2 in CO2 incubator while in group 2 oocytes matured in maturation media with 30 ng/ml melatonin supplementation. After 27 h of culture, nuclear maturation was observed in both groups using Hoechst dye. In experiment 2, 1,336 oocytes were randomly divided into two groups, viz. group 1 (641) wherein oocytes were matured in maturation media without melatonin while in group 2 (n=695) oocytes matured in maturation media with 30 ng/ml melatonin supplementation. After 27 h, oocytes of both groups were then subjected to in vitro fertilization.The rate of nuclear maturation in group 2 (30 ng/ml melatonin) was significantly higher than that of group 1 (control). Similarly, the cleavage rate and blastocyst formation from in vitro matured goat oocytes were significantly higher in group 2 than that of group 1. In conclusion, the result indicated that the supplementation of 30 ng/ml melatonin in maturation media improves the nuclear maturation and subsequent cleavage rates and blastocyst production from caprine oocytes.

The aim of the present study was to optimize the production of blastocyst for obtaining caprine embryonic stem cell-like cells. A total of 4372 cumulus oocyte complexes (COCs) were recovered by slicing the 1187 caprine ovaries and were matured in maturation media for 27 h in humidified atmosphere at 38.5°C with 5% CO2 in CO2 incubator. After 27 h of maturation, oocytes were denuded and were co-incubated with buck semen in fertilization medium (TALP medium + 8 mg/ml fatty acid free BSA and 50 μg/ml heparin) for 18 h. Good quality zygotes (2483) were selected and randomly divided into 2 groups (experiment 1), viz. Group 1 (1312) wherein the presumptive zygotes were cultured in RVCL while in Group 2 (1171) the presumptive zygotes were cultured in mCR2aa medium. The cleavage rate, blastocyst and hatched blastocyst production was significantly higher in Gr 1 (47.45±2.93, 10.13±1.31 and 3.90±1.13%) than Gr 2 (37.75±2.46, 4.20±0.93 and 1.66±0.72%). In experiment 2, after in-vitro fertilization, morula stage embryos and inner cell mass (ICM) from blastocyst and hatched blastocyst were used to isolate ES cell-like cells. Thus the results indicated that the RVCL medium is the best medium as far as the embryonic development up to blastocyst stage in comparison to mCR2aa media. Furthermore, the formation of putative embryonic stem cell colonies were higher from hatched blastocysts (91.6%) as compared to that of blastocysts (82.1%) and it was significantly higher than that from morulas (34.3%).