effect of biopsy during precompacted morula stage on post vitrification development blastocyst derived bovine embryos

ABSTRACT

Improvements on embryo micromanipulation techniques led to the use of embryo biopsy in commercial embryo transfer programs for genetic analysis of preimplantation bovine embryos. The aim of this study was to evaluate the quality of bovine blastocyst derived from embryos biopsied at different precompacted morulae stages by assessment of cryosurvivability of the resulting blastocysts. The in vitro produced bovine embryos were subjected to biopsy at days 2,3, and 4 post-isemination with different cell numbers [4 to 16-cells]. Embryo cell biopsy was carried out in a 100 micro l drop of H-SOF following pronase drilling by aspiration of one blastomrere. The biopsied embryos were then cutured in SOFFaaBSA co-cultured with oviduct cells-monolayer until blastocyst formation. The blastocysts were cryopreserved at room temperature after exposure of equilibration [glycerol 1.4 M for 5 min and then glycerol 1.4 M and ethylene glycol 3.6 M for 5 min] and vitrification solutions [3.4 M glycerol and 4.6 M ethylene glycol]. The blastocysts were loaded into the center of 0.25 ml straws separated by air bubbles from 2 columns of sucrose 0.5 M and plunged immediately into liquid nitrogen. There was no significant difference in cryosurvivability of vitrified-warmed blastocysts derived from biopsied embryos at different pre-compacted morula stages. The quality of biopsy derived blastocysts was identical to that of non-biopsy derived ones in terms of post vitrification survival and hatching rates. In conclusion there was no preference between different times of embryo biopsy at precompacted morula stages in term of cryosurvivability of biopsy derived bovine blastocysts