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KMID : 0900920000240040439
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Korean journal of Animal Reproduction
2000 Volume.24 No. 4 p.439 ~ p.449
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Expression of EGFP in Bovine Embryos after Intracytoplasmic Sperm Injection using Spermatozoa Co-cultured with Exogenous DNA
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Lee H.-C.
Uhm S.-J.
Ann S.-Y.
Chung H.-J.
Park H.-D.
Lee H.-T.
Chung K.-S.
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Abstract
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This study was to investigate the expression of transgene after co-injection of spermatozoon and EGFP gene into mature oocytes in cattle. From frozen semen, spermatozoa were treated by DTT with 0.03% Tween-20, freezing and thawing or 0.02% Triton X-100 to disrupt their plasma membranes. The sperm injected oocytes were co-cultured with bovine oviduct epithelial cells in CRlaa, and expression of EGFP in embryos were observbed under epifluorescent microscope. Two pronuclei were formed in oocytes injected with sperm treated by DTT (44.6%), DTT-Tween-20 (48.4%), DTT-freezing (44.4%) and DTT-Triton X-100 (42.9%). Cleavage and blastocyst formation rates of bovine oocytes which injected with sperm treated by DTT, DTT-Tween-20, DTT-freezing, and DTT-Triton X-100 were 49.1, 58.5, 43.9, and 48.4% and 10.2, 13.0, 6.8, and 6.5%, respectively. Although the most of embryos were showing mosaicism, embryos expressing EGFP gene were observed in all treated groups. Therefore, these results indicate that membrane disrupted sperm could interact with exogenous DNA, and that this procedure may be useful to introduce foreign gene into bovine oocytes.
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KEYWORD
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Exogenous DNA, Spermatozoa, ICSI, EGFP, Bovine
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