The use of green fluorescent protein (GFP) to select bovine embryos
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Institute of Genetics and Animal Breeding, Polish Academy of Sciences, Experimental Embryology Department, Jastrzębiec, 05-552 Wólka Kosowska, Poland
All Russian Research Institute for Farm Animal Genetics and Breeding, 55a Moskovskoe shosse, St. Petersburg-Pushkin, 189620 Russia
Institute of Genetics and Animal Breeding, Polish Academy of Sciences, Molecular Cytogenetics Department, Jastrzębiec, 05-552 Wólka Kosowska, Poland
Białystok Technical University, Faculty of Building and Environmental Engineering, Chair of Sanitary Biology and Biotechnology, Wiejska 45a, 15-351 Białystok, Poland
Publication date: 2003-01-02
J. Anim. Feed Sci. 2003;12(1):71–81
A major factor limiting the transgenesis in domestic animals is the inefficiency of maintaining large numbers of recipients carrying nontransgenic foetuses. The objectives of this study were: 1. to determine the influence of green fluorescent protein (GFP) construct injection on the development of bovine embryos, 2. to identify and select the GFP positive bovine embryos, and 3. to determine the rate of mosaicism in transgenic embryos. Cattle oocytes were matured and fertilised in vitro and zygotes were microinjected with pCX-EGFP construct consisting of CMV-IE enhancer, chicken β-actin promoter, cDNA of GFP (EGFP-732 bp) and rabbit β-globin polyadenylation sequences. Embryos from control (64) and microinjected (198) groups were cultured in vitro. After 168 h of culture, morula and blastocysts were observed in 39.06% of control and in 23.23% of injected group. We obtained three GFP positive embryos (1.51% of injected zygotes and 6.52% of morulae/blastocysts). One of them was 100, second 75 and third 25% GFP positive (66.7% of mosaicism). Use of gfp gene reporter to select bovine embryos is useful method to increase transgenic offspring, because GFP marker allows to choice only transgenic embryos and transfer them to recipients.