| 甘蓝型油菜乙酰辅酶A羧化酶3个亚基基因的克隆及其表达 |
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| 引用本文： | 武玉永,谭秀华,马立新.甘蓝型油菜乙酰辅酶A羧化酶3个亚基基因的克隆及其表达[J].安徽农业科学,2008,36(10):4002-4006. |
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| 作者姓名： | 武玉永 谭秀华 马立新 |
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| 作者单位： | 滨州医学院遗传学教研室,山东烟台,264003;湖北大学生命科学学院分子微生物学与基因工程实验室,湖北武汉,430062 |
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| 基金项目： | 湖北省自然科学基金项目(2003ABA118). |
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| 摘 要： | 目的]为构建乙酰辅酶A羧化酶的叶绿体表达载体奠定基础。方法]以从甘蓝型油菜叶片中提取的叶绿体基因组DNA为模板进行PCR扩增,克隆羧基转移酶β亚基基因。以从开花20~29 d后油菜幼胚中提取的总RNA为模板进行RT-PCR扩增,克隆生物素羧化酶和生物素羧基载体蛋白的cDNA序列。最后,乙酰辅酶A羧化酶的3个亚基基因被克隆到大肠杆菌表达载体中。结果]SDS-PAGE分析结果表明乙酰辅酶A羧化酶的3个亚基基因分别被克隆到大肠杆菌表达载体中。乙酰辅酶A羧化酶的3个亚基基因在大肠杆菌中的表达产物分别为76.6、44.0和81.5 kD,其融合蛋白分别占总菌体蛋白的12%、10%和6%。结论]不同品种的甘蓝型油菜生物素羧基载体蛋白cDNA序列存在较大差异。
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| 关 键 词： | 甘蓝型油菜 乙酰辅酶A羧化酶 基因克隆 基因表达 |
| 文章编号： | 0517-6611(2008)10-04002-05 |
| 修稿时间： | 2008年1月2日 |
Cloning of 3 Subunit Genes of Acetyl-CoA Carboxylase from Brassica napus L.and Their Expression in Escherichia coli |
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| WU Yu-yong.Cloning of 3 Subunit Genes of Acetyl-CoA Carboxylase from Brassica napus L.and Their Expression in Escherichia coli[J].Journal of Anhui Agricultural Sciences,2008,36(10):4002-4006. |
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| Authors: | WU Yu-yong |
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| Abstract: | Objective] The research aimed to lay the foundation for constructing the chloroplast expression vector of acetyl-CoA carboxylase.Method] The chloroplast genomic DNA extracted from the leaves of Brassica napus L.was taken as template for PCR amplification and the β-subunit gene of carboxyltransferase was cloned.Total RNA extracted from immature embryo of rape after flowering for 20～29 days was taken as template for RT-PCR amplification and the cDNA sequences of biotin carboxylase and biotin carboxyl carrier protein were cloned.Finally,3 subunit genes of acetyl-CoA carboxylase were cloned into the expression vector of E.coli.Result] The results of SDS-PAGE analysis showed that 3 subunit genes of acetyl-CoA carboxylase were successfully cloned into the expression vector of E.coli.The expression products of 3 subunit genes of acetyl-CoA carboxylase in E.coli were 76.6,44.0 and 81.5 kD resp.and their fusion proteins were 12%,10% and 6% of total bacterial protein resp.Conclusion] The cDNA sequence of biotin carboxyl carrier protein had greater difference among different varieties of B.napus L. |
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| Keywords: | Brassica napus L Acetyl-CoA carboxylase Gene cloning Gene expression |
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