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篇目详细内容 |
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【篇名】 |
Cloning and prokaryotic expression of TaE3 from wheat and preparation of antiserum |
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【刊名】 |
Frontiers of Agriculture in China |
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【刊名缩写】 |
Front. Agric. China |
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【ISSN】 |
1673-7334 |
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【EISSN】 |
1673-744X |
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【DOI】 |
10.1007/s11703-011-1146-7 |
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【出版社】 |
Higher Education Press and Springer-Verlag Berlin
Heidelberg |
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【出版年】 |
2011 |
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【卷期】 |
5
卷4期 |
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【页码】 |
437-442
页,共
6
页 |
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【作者】 |
Yunwei ZHANG;
Xiang GAO;
Shengfang HAN;
Dongmei WANG;
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【关键词】 |
E3 ubiquitin ligase (E3); wheat; prokaryotic expression; western blot analysis |
【摘要】 |
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The E3 ubiquitin ligase is a multi-functional protein that performs vital roles, particularly in various stress environment. To further understand the biological significance of E3 ubiquitin ligase gene from wheat (TaE3), total RNA was isolated from wheat leaves and then TaE3 gene was amplified by PCR after reverse transcription. The PCR product was cloned into PMD19-T vector to sequence subsequently. And then the recombinant expression vector (pET30a-GST-TaE3-His) was constructed and transformed into E. coli strain BL21 (DE3). SDS-PAGE analysis showed that the recombinant E. coli could express a proximate 43？kDa protein. TaE3 fusion protein was purified by Ni-NTA affinity chromatography from recombinant bacterial lysate and was used to immunize rabbit to produce polyclonal antibody. The titer and specificity of the anti-TaE3 antibody were successfully detected by indirect ELISA and western blot analysis. |
