Controversial aspect of using GFP as a marker for the production of transgenic cattle.
The objective of this study was to examine the feasibility of identification and selection of cattle embryos based on green fluorescence (GFP-positive) in order to obtain calves carrying an integrated transgene. The construct used (pbLGTNF-EGFP) contained the human tumor necrosis factor alpha (hTNFalpha) gene fused to the bovine beta-lactoglobulin promoter (bLG) in plasmid vector pCX-EGFP. In four experiments, 76 zygotes were injected; eight of them developed to the morulae/blastocysts stage of which only five were GFP positive (one of them 100%, one-50%, three- 25%). All of the GFP positive embryos were transferred to recipients. Two calves were born: one after transfer of the 100% GFP positive embryo and the other after transfer of one of the 25% GFP positive embryos. Both animals were healthy with normal weight when compared to two control calves. The integration of pbLGTNF-EGFP in the host genome could not be detected in either of the calves, suggesting that GFP is an unreliable marker for preimplantation screening of embryos.
Duszewska
A M
AM
Department of Experimental Embryology, Polish Academy of Sciences, Strzeszyńska 32, 60-479 Poznań, Poland. duszewskaanna@hotmail.com
Lipiński
D
D
Piliszek
A
A
Słomski
R
R
Pławski
A
A
Wojdan
J
J
Gawron
W
W
Juzwa
W
W
Zeyland
J
J
Wenta-Muchalska
E
E
Reklewski
Z
Z
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics*; D015415:Biomarkers; D001755:Blastocyst; D002417:Cattle / Q000235:genetics*; D004624:Embryo Transfer / Q000662:veterinary; D004622:Embryo, Mammalian / Q000166:cytology* / Q000378:metabolism; D005240:Feasibility Studies; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D007782:Lactoglobulins / Q000235:genetics; D008845:Microinjections / Q000662:veterinary; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000662:veterinary; D011993:Recombinant Fusion Proteins / Q000235:genetics; D019076:Transgenes / Q000235:genetics; D014180:Transplantation / Q000662:veterinary; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics
D015415:Biomarkers; D007782:Lactoglobulins; D011993:Recombinant Fusion Proteins; D014409:Tumor Necrosis Factor-alpha; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins