Eight-cell embryos collected from superovulated inbred strains and F1 hybrid mice were frozen by the microdrop technique developed in our laboratory. The technique based on pre-equilibration in medium with 10% glycerol, before transfer into vitrification solution, expel of embryos in 5 microliters to 20 microliters of vitrification solution directly into liquid nitrogen and thawing of microdrops in medium with 0.5 M sucrose was used. The behavior and morphological appearance of embryos during pre-freezing and post-thawing periods was documented. The efficiency of cryopreservation in microdrops was high, as documented by 90% to 100% of intact embryos after the freezing and thawing cycle. Furthermore, no zona pellucida damage was observed. The developmental potential of embryos frozen in microdrops was comparable with development of unfrozen embryos of the same genetic origin. After freezing and storage 83% to 93% of embryos developed to blastocysts and 73% to 92% embryos underwent "implantation" after 48 h and 96 h of in vitro culture, respectively.

• MeSH
• blastocysta cytologie   MeSH
• inbrední kmeny myší MeSH
• kryoprezervace metody   MeSH
• mikromanipulace MeSH
• myši MeSH
• techniky in vitro MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Zona-free zygotes, 2-cell, 4-cell, and 8-cell embryos, morulae, blastocysts and embryos exposed to biopsy and reconstruction were cryopreserved in microdrops of vitrification medium expelled directly into liquid nitrogen. While none of the zygotes and 2-cell embryos survived storage in liquid nitrogen, 54% 4-cell and 97% 8-cell embryos, 99% morulae, 88% blastocysts, 98% biopsied and 100% reconstructed embryos were intact one hour after thawing. Developmental potential of cryopreserved zona-free embryos evaluated after 30 h of in vitro culture was high. Of the frozen embryos, 49% 4-cell and 92% 8-cell embryos, 95% morulae, 79% blastocysts, 84% biopsied and 90% reconstructed embryos were capable of further cleavage. After the transfer of zona-free 8-cell embryos, morulae, blastocysts, biopsied and reconstructed embryos into day-1 recipients - 4 (16%), 14 (56%), 3 (12%), 0 and 8 (32%) live foetuses were recorded, respectively.

• MeSH
• embryo savčí * MeSH
• gastrula MeSH
• inbrední kmeny myší embryologie   MeSH
• kryoprezervace metody   MeSH
• morula MeSH
• myši MeSH
• zona pellucida MeSH
• zvířata MeSH
• zygota MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Two-cell embryos were collected from (C57BL/6 x CBA)F1 mice, microinjected with foreign DNA and transferred into temporary recipients. Intact and half 8-cell embryos recovered from oviducts 24 h later were frozen in microdrops of vitrification medium and stored in liquid nitrogen for 2 to 30 days. Development of frozen/thawed embryos was assessed by in vitro culture or transfer into day-1 recipients. From 1200 2-cell embryos 77% intact and 20% half embryos were obtained one hour after DNA injections. After transfer of 600 intact and 300 half embryos into temporary recipients, 77% intact and 73% half 8-cell embryos were recorded, respectively. Survival and in vitro development of frozen/thawed DNA-injected embryos were high, as 97% intact and 91% half 8-cell embryos developed to morulae or blastocysts. After transfer of 300 intact and 150 half 8-cell embryos into day-1 recipients, 99 (33%) and 19 (13%) young were born. None of the mice born from DNA-injected and cryopreserved embryos integrated injected genes.

• MeSH
• DNA genetika   MeSH
• embryo savčí * MeSH
• geneticky modifikovaná zvířata embryologie   MeSH
• inbrední kmeny myší MeSH
• injekce MeSH
• kryoprezervace metody   MeSH
• mikromanipulace MeSH
• myši MeSH
• těhotenství MeSH
• transfekce * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH

Transgenic mice were produced from DNA-injected embryos stored for 2 to 30 days in liquid nitrogen. Of the 500 zygotes collected from (C57BL/6 x CBA)F1 mice, 363 (73%) survived DNA injection into pronuclei and 246 (82%) morphologically normal 4- and 8-cell embryos were flushed from temporary recipients 48 h later. Of the 200 DNA-injected 8-cell embryos cryopreserved by vitrification in microdrops, 194 (97%) were recovered and 188 (94%) embryos were intact one hour after thawing. Of the 50 DNA-injected and frozen/thawed embryos, 48 (96%) developed to morulae or blastocysts within 30 h of in vitro culture. Transfer of 100 DNA-injected and cryopreserved 8-cell embryos into 20 day-1 recipients resulted in 47 young born. Two mice were transgenic.

• MeSH
• DNA genetika   MeSH
• embryo savčí fyziologie   MeSH
• erythropoetin genetika   MeSH
• exprese genu MeSH
• kryoprezervace * MeSH
• mikroinjekce MeSH
• míra přežití MeSH
• myši inbrední C57BL MeSH
• myši inbrední CBA MeSH
• myši transgenní genetika   MeSH
• myši MeSH
• přenos embrya MeSH
• transfekce * MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH
• erythropoetin MeSH