Knock-in of Enhanced Green Fluorescent Protein or/and Human Fibroblast Growth Factor 2 Gene into β-Casein Gene Locus in the Porcine Fibroblasts to Produce Therapeutic Protein

Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because...

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Published inAnimal bioscience Vol. 27; no. 11; pp. 1644 - 1651
Main Authors Lee, Sang Mi, Kim, Ji Woo, Jeong, Young-Hee, Kim, Se Eun, Kim, Yeong Ji, Moon, Seung Ju, Lee, Ji-Hye, Kim, Keun-Jung, Kim, Min-Kyu, Kang, Man-Jong
Format Journal Article
LanguageEnglish
Published Korea (South) Asian - Australasian Association of Animal Production Societies 01.11.2014
Asian-Australasian Association of Animal Production Societies (AAAP) and Korean Society of Animal Science and Technology (KSAST)
아세아·태평양축산학회
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ISSN1011-2367
1976-5517
2765-0189
1976-5517
2765-0235
DOI10.5713/ajas.2014.14222

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Summary:Transgenic animals have become important tools for the production of therapeutic proteins in the domestic animal. Production efficiencies of transgenic animals by conventional methods as microinjection and retrovirus vector methods are low, and the foreign gene expression levels are also low because of their random integration in the host genome. In this study, we investigated the homologous recombination on the porcine β-casein gene locus using a knock-in vector for the β-casein gene locus. We developed the knock-in vector on the porcine β-casein gene locus and isolated knock-in fibroblast for nuclear transfer. The knock-in vector consisted of the neomycin resistance gene (neo) as a positive selectable marker gene, diphtheria toxin-A gene as negative selection marker, and 5' arm and 3' arm from the porcine β-casein gene. The secretion of enhanced green fluorescent protein (EGFP) was more easily detected in the cell culture media than it was by western blot analysis of cell extract of the HC11 mouse mammary epithelial cells transfected with EGFP knock-in vector. These results indicated that a knock-in system using β-casein gene induced high expression of transgene by the gene regulatory sequence of endogenous β-casein gene. These fibroblasts may be used to produce transgenic pigs for the production of therapeutic proteins via the mammary glands.
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Department of Animal Science, College of Agriculture and Life Science, Chonnam National University, Gwangju 500-757, Korea
G704-001112.2014.27.11.020
ISSN:1011-2367
1976-5517
2765-0189
1976-5517
2765-0235
DOI:10.5713/ajas.2014.14222