油菜天然抗性巨嗜蛋白（BnNRAMP）的筛选与基因克隆
毕业论文关键词：BnNRAMP1b；油菜；镉离子转运；生物信息学；基因克隆
The Characterization and Representative Gene Construction of Natural Resistance-Associated Macrophage Proteins in Brassica napus
Abstract：Cadmium causes a serious environmental problem all over the world as a major element in heavy-metal pollution from agricultural and industrial emissions. The consequent chronic cadmium poisoning also leads to great damage to human health. In many areas in China, it becomes extremely important to develop chemical or biological technics for remediation of contaminated soils, in order to solve all kinds of heavy-metal environment issues. Particularly, great potentials were discovered from phytoremediation strategy by utilizing crops with wide cultivating areas and short growth cycles because of low cost and less side-effects. As a result, Brassica napus has become one of the most popular materials due to its remarkable bioaccumulation ability of cadmium. The previous investigations in our lab have demonstrated that B. napus changes a lot of physiological features with regulations of NRAMP family under cadmium stress. However, the detailed mechanism and the molecular functions of NRAMP family in B. napus remain unclear. Therefore, this article concentrates on the evolutional bioinformatic analysis, transcriptional changes of NRAMP family under the cadmium stress and the cloning of BnNRAMP1b gene, which provides materials for future validations of cadmium transport functions of BnNRAMP1b. It is also a part of theoretical evidence for further investigations of B. napus’s application value in phytoremediation strategy.
Key words: BnNRAMP1; Brassica napus; Cadmium Transport; Bioinformatics; Cloning
目 录
摘要3
关键词3
Abstract。3
Key words3
引言（或绪论）。3
1 材料与方法。5
1.1 材料 。5
1.2 方法 。5
1.2.1 BnNRAMP基因家族筛选办法。5
1.2.2 Brassica napus cDNA文库构建5
1.2.3油菜基因表达数据库6
1.2.4 BnNRAMP1基因克隆。6
1.2.5 BnNRAMP1b克隆载体构建。7
1.2.6 BnNRAMP1b转导大肠杆菌。7
1.2.7 BnNRAMP1b/pEASY质粒提取。8
1.2.8 构建酵母pYES2蛋白表达载体。8
1.2.9 鲑鱼精DNA法酵母转化8
2 结果与分析8
2.1 BnNRAMP家族蛋白的筛选8
2.1.1 BnNRAMP同源蛋白表。8
2.1.2 BnNRAMP家族进化分析9
2.1.3 BnNRAMP同源蛋白结构域分析10
2.1.4 BnNRAMP家族在镉胁迫条件下的表达。11
2.2 BnNRAMP1b基因克隆12
2.3 BnNRAMP1b蛋白镉离子转运功能的验证13
2.3.1构建ycf-/-/BnNRAMP1b+酵母模型。13
3 讨论。14
致谢15
参考文献。15
油菜天然抗性巨嗜蛋白（BnNRAMP）的筛选与基因克隆