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KMID : 0900920030270030259
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Korean journal of Animal Reproduction
2003 Volume.27 No. 3 p.259 ~ p.268
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Inducible Expression of the Lactadherin Gene with a Reverse Tetracycline-Regulated Retroviral Vector System
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Kim Teo-An
Park Jae-Bok
Oh Hoon-Kyu
Kwon Mo-Seon
Park Chang-Sik
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Abstract
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Lactadherin (formerly known as BA46), a major glycoprotein of the human milk fat globule membrane, is abundant in human breast milk and breast carcinoma cells and is known to prevent symptomatic rotavirus infections. In this study, we tried to transfer the human lactadherin gene to the Chinese Hamster Ovary (CHO) cells using retrovirus vector system and tested inducible expression of the gene under the tetracycline-controllable promoter. At first, tetracycline-mediated inducibility was tested using E.coli LacZ marker gene. NIH3T3 cells co-infected with RevTet-On and RevTRE-LacZ retrovirus vectors showed that the cells responded to doxycycline (a derivative of tetracycline) in a dose-dependent manner, and prominent induction of the lacZ gene expression was observed from 1 mutextrm{g}/ml of doxycycline concentration. Based on the results of the pilot experiment, inductional expression of the human lactadherin gene was conducted using RevTet-On and RevTRE-Ltd retrovirus vectors. Analysis with the RT-PCR demonstrated successful inductional expression of the lactadherin gene in the Chinese Hamster Ovary (CHO) cells. Considering that constitutive overexpression of the exogenous genes in the target cells causes serious physiological imbalance, the results obtained in this study will be very useful especially in the studies of gene therapy and transgenic animal production.
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KEYWORD
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Ltd gene, Inducible expression, Doxycycline, CHO cell, RT-PCR
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