油菜天然抗性巨嗜蛋白（BnNRAMP）的筛选与基因克隆

摘要：镉作为一种农业工业生产排放的主要重金属元素，在全球范围内导致了非常严重的环境问题。其带来的慢性镉中毒也严重危害着人类健康。在中国许多地区，为解决各种重金属污染问题，研究修复污染土壤及水源的化学、生物学技术变得极为重要。其中，因其成本低、副作用小等优点，利用种植面积大、生长周期短的农作物进行土壤修复的技术拥有极大应用潜力。对此，具有极强的镉元素富集作用甘蓝型油菜成为了首选的植物素材之一。我们实验室前期工作发现，油菜在镉胁迫生长条件下，伴随着NRAMP基因的表达变化，产生了一系列的生理变化。然而，其植株具体的应对机理以及油菜NRAMP家族的分子学功能尚不明确。本文针对这一空白，分析鉴定了油菜NRAMP家族蛋白的进化信息及其在镉胁迫条件下的表达变化，成功克隆出了BnNRAMP1b基因，为证实BnNRAMP1b对镉离子的转运功能提供了材料，也为进一步探究证实甘蓝型油菜修复镉污染土壤的应用价值提供了理论依据。25630
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）的筛选与基因克隆油菜天然抗性巨嗜蛋白（BnNRAMP）的筛选与基因克隆:http://www.751com.cn/shengwu/lunwen_19510.html