MICROINJECTION OF BOVINE EMBRYOS WITH A FOREIGN GENE AND ITS DETECTION AT THE BLASTOCYST STAGE

This study was designed to measure the survival and development of pro nuclear stage bovine zygotes after microinjection with DNA and either culture in vitro or incubation in the sheep oviduct and to determine t he percentage of embryos that retain exogenous DNA at the blastocyst s tage. In vitro matured and fertilized oocytes were examined for pronuc lear development 18 to 20 h after coincubation with sperm. An ovine so matotropin gene construct was microinjected into ova with visible pron uclei. Microinjected ova and unmanipulated ova from the same in vitro derived pool were cocultured with oviductal epithelial cells or incuba ted in the sheep oviduct and cocultured, respectively, for 7 d. Blasto cysts were subjected to polymerase chain reaction analysis for detecti on of exogenous DNA. The percentage of cleaved embryos that formed bla stocysts was similar for microinjected and unmanipulated cultured ova. The percentage of blastocysts recovered from the sheep oviduct that h atched was higher than for blastocysts that developed in coculture. Fo urteen of 26 (54%) blastocysts that developed from microinjected ova w ere positive for the exogenous DNA by polymerase chain reaction. The i mplications are discussed for polymerase chain reaction detection of t he exogenous gene construct in a relatively high proportion of microin jected blastocysts.