摘要：【目的】从‘巨峰’葡萄中克隆白藜芦醇合酶基因Rs，对其序列进行结构及功能分析，并鉴定其在果实不同发育时期的时空表达特异性。【方法】利用CTAB法提取葡萄总RNA，采用RT-PCR技术克隆Rs基因，借助qRT-PCR技术对其表达量进行分析。利用生物信息学工具对其核酸及蛋白质序列进行分析。【结果】克隆出的Rs基因cDNA序列全长1247bp，有一个长度1179bp的开放阅读框（ORF）；该基因含有查耳酮和二苯乙烯合酶活性位点以及完整的茋合酶家族特征位点。qRT-PCR检测结果表明，Rs基因在‘巨峰’葡萄花后25d的青皮中相对表达量最高。【结论】本研究克隆出了Rs基因，并对其进行序列结构分析，鉴定了其时空表达特异性，为进一步认识Rs基因在白藜芦醇代谢过程中的作用奠定了基础。28055
毕业论文关键词：葡萄；Rs基因；克隆；白藜芦醇
Cloning, Sequence Analysis and Characterization of Rs Gene in Vitis vinifera
Abstract：【Objective】To clone the resveratrol synthase gene Rs from Vitis vinifera cv  Kyoho, and analyze its sequence and evolution and identify the specificity of the protein in the different developmental stages of fruit  【Method】The total RNA was extracted by CTAB method  The Rs gene was cloned by RT-PCR  The expression of Rs gene was analyzed by qRT-PCR  The sequence of nucleic acid and protein were analyzed by using bioinformatics tools  【Result】The cDNA sequence of the cloned Rs gene was 1247bp in length, and there was an open reading frame (ORF) with a length of 1179bp  The gene contained chalcone and stilbene synthase active sites and intact stacclase family Point  The results of qRT-PCR showed that the relative expression of Rs gene was the highest in green peel of 25 days after flowering  【Conclusion】The Rs gene was cloned and analyzed, and the specificity of spatiotemporal expression was identified  It laid a foundation for further understanding the role of Rs gene in the process of resveratrol metabolism
Key words: Vitis vinifera L , Rs Gene, Clone, Resveratrol
目  录
摘要    1
关键词    1
Abstract     1
Key words    1
1  材料与方法    2
1．1  材料    2
1．1．1  植物材料    2
1．1．2  菌株和载体    2
1．1．3  酶及生化试剂    2
1．1．4  引物与测序    2
1．1．5  培养基    2
1．1．6  主要仪器与设备    2
1．1．7  生物信息学分析    3
1．2  方法    3
1．2．1  葡萄基因组总RNA的提取及cDNA合成    3
1．2．2  目的基因的分子克隆    3
1．2．3  基因的生物信息学分析    3
1．2．4  基因表达特异性分析    3
1．2．5  葡萄皮愈伤组织的诱导与培养    4
1．2．6  细菌转化方法    4
2  结果与分析    4
2．1  葡萄基因组总RNA的提取及cDNA合成    4
2．2  目的基因的克隆    5
2．2．1  基因克隆结果    5
2．2．2  遗传转化结果    5
2．2．3  双酶切结果    6
2．3  基因的生物信息学分析    6
2．3．1  Motif分析    6
2．3．2  基因结构分析    7
2．3．3  蛋白质理化性质及氨基酸组成分析    7
2．3．4  二级结构预测    7 葡萄Rs基因的克隆序列分析及时空表达特征鉴定:http://www.751com.cn/shengwu/lunwen_22765.html