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目的构建红花查耳酮异构酶(CHI)基因的植物表达载体并在拟南芥中进行超表达验证该基因的功能。方法将已经分离的红花CHI基因作为目的基因,在其两端引入Bam H I和Eco R I酶切位点,构建含有35S启动子的植物超表达载体p BASTA-CHI,通过Flora-dip法将其转化到拟南芥中,并对转基因拟南芥T2代植株进行PCR和总黄酮量检测。结果转基因拟南芥T2代植株的PCR检测,红花CHI基因已经初步整合到拟南芥基因组中,黄酮量检测表明,转红花CHI基因的拟南芥比野生型拟南芥中的黄酮量有所提高,最高提高到2.3倍。结论成功构建了含有红花CHI基因的植物表达载体,并在拟南芥中进行超表达,获得了转基因拟南芥T2植株。
Objective To construct plant expression vector of chalcone isomerase (CHI) gene and validate the function of this gene in Arabidopsis. Methods The chimpanzee CHI gene was cloned and inserted into Bam HI and Eco RI restriction sites at both ends to construct a plant over expression vector p BASTA-CHI containing 35S promoter. Transformed into Arabidopsis thaliana, and T2 generation of transgenic Arabidopsis plants PCR and total flavonoids detection. Results PCR analysis of T2 generation transgenic Arabidopsis thaliana plants showed that the safflower CHI gene had been initially integrated into Arabidopsis thaliana genome. Flavonoids detection showed that the content of flavonoids in Arabidopsis thaliana transformed with CHI gene was higher than that in wild-type Arabidopsis thaliana Increased, up to 2.3 times. Conclusion The plant expression vector containing safflower CHI gene was successfully constructed and overexpressed in Arabidopsis. Transgenic Arabidopsis T2 plants were obtained.