This study was carried out to investigate the effects of cryoprotectants, warming solution and removal of lipid on open pulled straw vitrification (OPS) method of porcine embryos produced by nuclear transfer (NT) of fetal fibroblasts. All solutions used during vitrification were prepared with holding medium consisting of 25 mM Hepes buffered TCM199 medium containing 20% fetal bovine serum (FBS) at 38.5℃. The blastocysts derived from NT with or without lipid were vitrified in each medium of different concentrations of dimethyl sulfoxide (DMSO) and ethylene glycol (EG). Also, blastocysts after cryopreservation were warmed into different concentrations of sucrose in warming solution. The optimal concentrations of cryoprotectants in vitrification solution were 10% DMSO + 10% EG in vitrification solution 1 (VS1) and 20% DMSO + 20% EG in vitrification solution 2 (VS2). The optimal concentrations of sucrose were 0.3 M sucrose in warming solution 1 (WS1) and 0.15 M sucrose in warming solution 2 (WS2). Lipid removal from oocytes before NT enhanced the viability of NT embryos after vitrification. Our results show that use of the OPS method in conjunction with lipid removal provides effective cryopreservation of porcine nuclear transfer embryos.

본 연구는 미성숙, 성숙 단계의 돼지 난포란이 유리화 동결에 의한 동결보존이 가능한지를 조사 하고자 실시하였다. 난포란은 세포질 내 지방구를 분극시키기 위해 원심분리를 실시하였고, 미세조작기를 이용하여 지방구를 제거하였다. 돼지 난포란을 CB 처리하여 원심분리 후 지방구를 제거한 지방제거구(Delipated), CB 처리 후 원심분리만 하여 지방구를 분극시킨 원심분리구(Centrifuged), 아무처리도 하지 않은 대조구(Control)를 EM grid