| 羽衣甘蓝S_(13-b)位点受体激酶基因的克隆及激酶结构域的原核表达 |
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| 引用本文： | 蓝兴国,杨佳,王艳红,李玉花. 羽衣甘蓝S_(13-b)位点受体激酶基因的克隆及激酶结构域的原核表达[J]. 生物技术通讯, 2010, 21(2): 210-212. DOI: 10.3969/j.issn.1009-0002.2010.02.015 |
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| 作者姓名： | 蓝兴国 杨佳 王艳红 李玉花 |
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| 作者单位： | 东北林业大学,生命科学学院,黑龙江,哈尔滨,150040 |
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| 摘 要： | 目的:分离羽衣甘蓝S13-b位点受体激酶(SRK13-b)基因并进行序列及结构域分析,构建SRK13-b结构域的原核表达载体并进行重组蛋白质的原核表达和纯化。方法:提取羽衣甘蓝S13-bS13-b自交不亲和系花期柱头的RNA,用RT-PCR法分离SRK13-b基因;将编码SRK13-b激酶结构域的序列插入大肠杆菌表达载体pET-14b中,构建原核表达质粒pET-SRK13-bCT,转化大肠杆菌BL21(DE3)pLysS菌株,经0.1mmol/LIPTG诱导,用Ni-NTA亲和层析柱对SRK13-b激酶结构域蛋白进行纯化。结果:分离获得羽衣甘蓝SRK13-b基因的长度为2571bp,编码856个氨基酸,GenBank收录号为EU180597;对SRK13-b激酶结构域蛋白进行诱导表达及纯化,SDS-PAGE显示相对分子质量约43×103的蛋白质特异表达,对表达产物进行分离纯化,获得了SRK13-b激酶结构域的融合蛋白。结论:羽衣甘蓝SRK13-b基因的克隆及激酶结构域的原核表达,为研究SRK的功能及自交不亲和性奠定了基础。
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| 关 键 词： | 羽衣甘蓝 自交不亲和 S13-b位点受体激酶 原核表达 纯化 |
Cloning of SRK_(13-b) Gene of Ornamental Kale (Brassica oleracea var.acephala) and Prokaryotic Expression of Kinase Domain of SRK_(13-b) |
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| LAN Xing-Guo,YANG Jia,WANG Yan-Hong,LI Yu-Hua. Cloning of SRK_(13-b) Gene of Ornamental Kale (Brassica oleracea var.acephala) and Prokaryotic Expression of Kinase Domain of SRK_(13-b)[J]. Letters in Biotechnology, 2010, 21(2): 210-212. DOI: 10.3969/j.issn.1009-0002.2010.02.015 |
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| Authors: | LAN Xing-Guo YANG Jia WANG Yan-Hong LI Yu-Hua |
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| Affiliation: | LAN Xing-Guo,YANG Jia,WANG Yan-Hong,LI Yu-Hua College of Life Sciences,Northeast Forestry University,Harbin 150040,China |
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| Abstract: | Objective:To isolate the S13-b-locus receptor kinase(SRK13-b) gene of ornamental kale(Brassica oleracea var.acephala),and obtain the purified recombinant fusion protein SRK13-bCT expressed in Escherichia coli.Methods:Total RNA was extracted from the flowering stigma of ornamental kale S13-b homozygotes.The full-length coding region of SRK13-b was isolated by using RT-PCR.The cytoplasmic kinase coding region of SRK13-b was inserted into an expression vector,pET-14b,and generated recombinant plasmid pET-SRK13... |
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| Keywords: | Brassica oleracea var.acephala self-incompatibility S13-b-locus receptor kinase prokaryotic expression purification |
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