Nutritional impact on gene expression and competence of oocytes used to support embryo development and livebirth by cloning procedures in goats.

Changes in the nutritional plan have been shown to affect oocyte quality, crucial to oocyte donors animals used in cloning. This study aimed to evaluate the impact of diets with increasing nutritional levels (maintenance diet=M; 1.3M; 1.6M; 1.9M) fed to goats for four weeks on follicular fluid composition, gene expression and oocyte competence used to cloning in goats. Donor females were superovulated for the retrieval of matured oocytes and physical measurements reported. After four weeks, groups receiving diets above maintenance increased thickness of subcutaneous adipose tissue and body weight, with higher values in 1.9M Group (P<0.05). Treatments did not affect follicular density, number of aspirated follicles, retrieved and matured oocytes. Animals from 1.3M group had lower (P<0.05) maturation rate (44.0%) and number of viable oocytes (65.3%) than M (68.8%) and 1.9M (76.0%). Follicular fluid glucose concentrations increased with nutritional levels (P=0.010), with a difference (P<0.05) between groups 1.9M (11.4±2.6mg/dL) and M (2.6±0.5mg/dL). The diet did not affect the expression of GDF9, BMP15, and BAX genes in oocytes, but BCL2 and apoptotic index were significantly higher (P<0.05) in the 1.3M and 1.6M groups than the other groups. Following the transfer of cloned embryos, one fetus was born live of a twin pregnancy in the 1.9M Group. The association between energy intake and oocyte quality suggests better nutritional use by oocytes when the maximum flow was used (1.9M), but the optimal feeding level in cloning still needs refinement.

A fast and simple method for the polymerase chain reaction-based sexing of livestock embryos.

Embryo sexing is a powerful tool for livestock producers because it allows them to manage their breeding stocks more effectively. However, the cost of supplies and reagents, and the need for trained professionals to biopsy embryos by micromanipulation restrict the worldwide use of the technology to a limited number of specialized groups. The aim of this study was to couple a fast and inexpensive DNA extraction protocol with a practical biopsy approach to create a simple, quick, effective, and dependable embryo sexing procedure. From a total of 1847 sheep and cattle whole embryos or embryo biopsies, the sexing efficiency was 100% for embryo biopsies, 98% for sheep embryos, and 90.2% for cattle embryos. We used a primer pair that was common to both species and only 10% of the total extracted DNA. The whole protocol takes only 2 h to perform, which suggests that the proposed procedure can be readily applied to field conditions. Moreover, in addition to embryo sexing, the procedure can be used for further analyses, such as genotyping and molecular diagnosis in preimplantation embryos.