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| Immune BlotAnalysis on Expression of the Mammalian Target of Rapamycin in Goat Fetal Fibroblasts with Recombinant Polyclonal Antibody |
| LIANGYan, WANGXiao-jing, YANGJiao-fu, HAOXi-yan, BAYin-ma, CHENXian-wei WANGZhi-gang |
| College of Life Science, Inner Mongolia University, Hohhot 010021, P.R.China |
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摘要 To detect the mammalian target of rapamycin (mTOR) expressed in Cashmere goat fetal fibroblasts (GFb), mTOR gene was cloned from Inner Mongolia Cashmere goat (Capra hircus) and expressed in Escherichia coli followed by immunizing mice with the purified recombinant protein as an immunogen to produce the anti-goat mTOR recombinant polyclonal antibody. Antiserum was collected from the immunized mice after the fifth immunization and its titer was determined with enzyme-linked immunosorbent assay (ELISA). The results showed that the recombinant polyclonal antibody had a titer 1:200 000 and could react with the mTOR expressed in GFb cells with a specific and sensitive affinity. Western blot showed that mTOR expression and phospho-mTOR (Ser 2448) activity were inhibited when GFb cells were treated with CCI-779, an mTOR specific inhibitor.
Abstract To detect the mammalian target of rapamycin (mTOR) expressed in Cashmere goat fetal fibroblasts (GFb), mTOR gene was cloned from Inner Mongolia Cashmere goat (Capra hircus) and expressed in Escherichia coli followed by immunizing mice with the purified recombinant protein as an immunogen to produce the anti-goat mTOR recombinant polyclonal antibody. Antiserum was collected from the immunized mice after the fifth immunization and its titer was determined with enzyme-linked immunosorbent assay (ELISA). The results showed that the recombinant polyclonal antibody had a titer 1:200 000 and could react with the mTOR expressed in GFb cells with a specific and sensitive affinity. Western blot showed that mTOR expression and phospho-mTOR (Ser 2448) activity were inhibited when GFb cells were treated with CCI-779, an mTOR specific inhibitor.
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Received: 11 February 2011
Accepted:
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| Fund: This work supported by the National Natural Science Foundation of China (30860191), the Graduate Student Research Projects of Inner Mongolia University and Major Projects for New Varieties of Genetically Modified Organisms of China (2008ZX08008-002). |
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Corresponding Authors:
WANG Zhi-gang, Mobile: 13190603899, E-mail: lswzg@imu.edu.cn
E-mail: lswzg@imu.edu.cn
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| About author: LIANG Yan, Mobile: 15847152620, E-mail: xixils@163.com |
Cite this article:
LIANGYan , WANGXiao-jing , YANGJiao-fu , HAOXi-yan , BAYin-ma , CHENXian-wei WANGZhi-gang .
2012.
Immune BlotAnalysis on Expression of the Mammalian Target of Rapamycin in Goat Fetal Fibroblasts with Recombinant Polyclonal Antibody. Journal of Integrative Agriculture, 12(6): 1002-1008.
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