Effects of Time of Deoxyribonucleic Acid Microinjection on Gene Detection and In Vitro Development of Bovine Embryos

• Published in: Journal of dairy science Vol. 77; no. 3; pp. 718 - 724
• Main Authors: Sparks, A.E.T., Canseco, R.S., Russell, C.G., Johnson, J.L., Moll, H.D., Velander, W.H., Gwazdauskas, F.C.
• Format: Journal Article
• Language: English
• Published: Savoy, IL Elsevier Inc 01.03.1994; Am Dairy Sci Assoc; American Dairy Science Association
• Subjects: ADN; Animals; Animals, Genetically Modified - embryology; Base Sequence; Biological and medical sciences; Biotechnology; Blastocyst - physiology; bovine embryo culture; Cattle; CULTIVO DE EMBRIONES; CULTURE D'EMBRYON; Culture Techniques; deoxyribonucleic acid microinjection; DESARROLLO EMBRIONARIO; DEVELOPPEMENT EMBRYONNAIRE; DNA - administration & dosage; Embryonic and Fetal Development; EXPERIMENTACION IN VITRO; EXPERIMENTATION IN VITRO; Female; Fundamental and applied biological sciences. Psychology; GENE; Gene Transfer Techniques; GENES; Genetic engineering; Genetic technics; Methods. Procedures. Technologies; Microinjections; Molecular Sequence Data; Morula - physiology; Polymerase Chain Reaction; SUPEROVULACION; SUPEROVULATION; Time Factors; Transgenic animals; Transgenic animals and transgenic plants; VACAS LECHERAS; VACHE LAITIERE; OEC; deoxyribonucleic acid microinjection; bovine embryo culture; PCR; Performance evaluation; Embryonic development; Genetic selection; Bovine; Transgenic animal; Genetic transfer; Vertebrata; Mammalia; Microinjection; Embryo culture; Artiodactyla; Kinetics; Ungulata
• DOI: 10.3168/jds.S0022-0302(94)77005-3
• ISSN: 0022-0302

In vivo fertilized embryos were surgically collected from superovulated dairy cows to evaluate microinjection on embryo development and utilized the polymerase chain reaction technique for selection of transgenic embryos. Seventy-two percent of the embryos with visible pronuclei or nuclei were microinjected with DNA, and the remaining 28% served as uninjected controls. All embryos were cocultured with bovine oviductal epithelial cells. Mean final development scores of embryos within the same initial cell stage at collection were unaffected by microinjection. After 144h of culture, 45% of the microinjected embryos developed to the morula or blastocyst stage. The transgene was detected in 50, 10, and 9% of demimorulae from embryos microinjected at the 1-, 2-, and 4-cell stages. Frequency of transgene detection was higher in morulae from 1-cell embryos than in morulae from 2- and 4-cell embryos. Use of in vitro coculture, embryo bisection, and polymerase chain reaction technique facilitated selection of bovine embryos that carried the transgene.