Production of transgenic mice from DNA-injected embryos cryopreserved by vitrification in microdrops

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.