Microsurgical separation of deconserved embryos

Abstract

At the current level of development of cell engineering methods in cattle breeding, the accelerated production of monozygotic twins is acquiring important scientific and economic significance. This will make it possible to determine the effectiveness of breeding methods in assessing genetic progress in a population over a number of generations. The method of embryonic cloning by microsurgical separation of embryos is close to solving this problem. Of particular interest is the long-term preservation of demi-embryos to obtain genetic analogues. But the possibilities of preserving demi-embryos at low temperatures are limited, since existing methods are multi-stage and extremely difficult to implement in practice. The search for applied methods of microsurgery in combination with long-term preservation of demi-embryos is an urgent task. Early studies have reported micromanipulation of frozen-thawed embryos. In 1984, in experiments by S. Willadsen & R. Godke, after microsurgery of deconserved embryos, the survival rate was 54.2%. Data on survival rate are not provided. No other data was found, and therefore we conducted studies to determine the influence of the processes of cryo- and deconservation, and dissection of embryos on the survival and ability of demi-embryos to develop totipotently after transplantation. The research was carried out in 2022 at the Regional Research and Production Center for the Reproduction of Farm Animals of Kalmyk State University named after B.B. Gorodovikov. The efficiency of dissection of intact embryos was 86.1% - after separation of 61 embryos, 105 suitable demi-embryos were obtained out of 122 possible, which was 5.7% higher than the same figure for dividing deconserved embryos - 80.4% (37 suitable demi-embryos out of 46 possible after separation 23 embryos). The survival rate of demi-embryos as the ability to nidate in the recipient's uterus in intact and depreserved demi-embryos differed within acceptable limits - 52.2% when transplanting 1 intact demi-embryo (12 pregnant out of 23 recipients) and 65.9% when transplanting 2 demi-embryos (27 pregnant from 41). Similar indicators for transplantation of deconserved demi-embryos were respectively: 42.8 (3 pregnant recipients out of 7 transplanted) and 60.0% (9 pregnant recipients out of 15).