【作者】 王治涛；

【导师】 徐莺；

【作者基本信息】 四川大学 ， 植物学， 2006， 硕士

【摘要】 以等质量比的不同发育阶段的麻疯树（Jatropha curcas L．）胚乳为材料，提取纯化mRNA，使用Stratagene公司生产的cDNA Synthesis Kit、Gigapack Ⅲ Gold Packaging Extract，用Uni-ZAP XR载体构建麻疯树胚乳cDNA文库。初级文库滴度为1.5×10⁶Pfu／ml，扩增文库的滴度为1.0×10¹⁴pfu／mL，重组率约为99.7％，插入片段大小范围经PCR及测序检测为0.4kb～4kb，平均约1kb。随机挑耳义10个cDNA插入片段进行测序，利用生物信息学方法，与其它生物已知的基因或蛋白质序列做全长性与同源性比较分析，结果表明文库中的cDNA插入片段的完整性较好，全长基因的比例达到20％，可以用于全长基因的筛选。根据测得cDNA序列推测出的蛋白质序列种类主要包括贮存蛋白相关、转录表达调控相关、脂肪酸合成相关及信号传导相关等同源基因序列，具体有金属硫因蛋白、翻译延长因子、MADS—Box等基因的类似物。参与贮存蛋白相关基因的比例是20％，转录因子相关基因的比例是6％，脂肪酸合成代谢的比例是4％，信号传导相关基因的比例是12％，氨基酸代谢相关基因的比例是2％。编码功能基因的类型与分布情况同Joseph A．White等构建的蓖麻胚乳cDNA文库中的基因的类型与分布情况基本一致，个别类型如贮存蛋白合成相关基因的比例相比较低，信号传导与转录引子相关基因的比例较高。 磷酸烯醇式丙酮酸羧化酶（PEPCase，EC 4.1.1.31）是一种CO₂固定酶，它催化从磷酸烯醇式丙酮酸（PEP）和碳酸氢酐（CA）生成草酰乙酸（OAA）的反应，在能量和生物合成代谢中主要发挥多种作用。麻疯树PEPC基因的克隆从双子叶植物与单子叶植物中同源性高的PEPC基因的序列比较分析开始，更多还原

【Abstract】 Equal weight endosperm of Jatropha curcas （Linnaeus） at different development stage was used as experimental materials. Total RNA is extracted and purified. Jalropha curcas L. endosperm cDNA library was constructed using cDNA Synthesis Kit, Gigapack III Gold Packaging Extract and Uni-ZAP XR vector. A primary library of 1.5×10^<sup>6Pfu/ml was obtained. An amplified library of 1.5×10¹⁴Pfu/ml was obtained. The recombinant efficiency was 99.7%. The inserted fractions sizes ranged from 0.4kb to 4kb according to the PCR results and sequenced analysis, with an average value of 1kb. After sequenced 10 randomly selected cDNA, Homologous and full-length analysis with other genes indicated that all of picked clones were integrated by using bio-informational methods. The percentage of full-length genes was 20%. This suggests that this library might be suitable to be used to screen full-length gene. These sequences mainly involved in the process of storage protein metabolism, gene expression and their regulation, fatty acid metabolism related genes and signal transduction related genes, including metallothionein-like protein, translation elongation factor and MADS-box gene analog et al. The percentage of storage protein related genes was 20%. The percentage of transcriptional factor related genes was 6%. The percentage of fatty更多还原