灯盏花MYB基因克隆及其荧光表达载体的构建

发布时间：2018-07-24 16:31

【摘要】：目的克隆灯盏花MYB基因的全长序列,为解析灯盏花MYB基因的功能奠定基础。方法根据灯盏花转录组的相关信息,利用RACE方法从灯盏花中克隆到一个可能参与灯盏花乙素合成的MYB基因,在对其cDNA序列、核苷酸序列的相似性、理化性质、疏水性、跨膜结构、二级结构及三级结构进行分析预测的基础上,对其进行多序列比对并构建系统树。同时,还构建了该基因与绿色荧光蛋白的融合表达载体,并进行了初步转化研究。结果克隆获得灯盏花MYB基因,命名为ebMYB06,其开放阅读框为783 bp,编码260个氨基酸残基,相对分子质量为63 800,理论等电点(p I)为5.18,属稳定蛋白。其蛋白二级结构主要由无规卷曲、α-螺旋和β-折叠构成。根据灯盏花MYB与拟南芥MYB(AtMYB)的系统树比对分析结果,发现ebMYB基因与拟南芥中的AtMYB4、7、32、6、8和AtMYB11、12、111 2亚群的基因聚类,推测所克隆基因在结构或功能上,可能与这两组具有共同性。实验还表明所构建的表达可用于灯盏花的高效转化。结论首次从灯盏花中克隆到可能参与其苯丙烷代谢或环境响应调控的MYB基因。
[Abstract]:Objective to clone the full-length sequence of MYB gene from Erigeron breviscapus and to lay a foundation for analyzing the function of MYB gene in Erigeron breviscapus. Methods according to the information of breviscapine transcriptome, a MYB gene was cloned from Erigeron breviscapus by RACE method. Its cDNA sequence, nucleotide sequence similarity, physicochemical properties and hydrophobicity were analyzed. Based on the analysis and prediction of the transmembrane structure, secondary structure and tertiary structure, the multi-sequence alignment is carried out and the system tree is constructed. At the same time, the fusion expression vector of the gene and green fluorescent protein was constructed, and the preliminary transformation was carried out. Results the MYB gene of Erigeron breviscapus was cloned and named ebMYB06.The open reading frame was 783 BP, encoding 260 amino acid residues. The relative molecular weight was 63 800 and the theoretical isoelectric point (p I) was 5.18, which was a stable protein. The protein secondary structure is mainly composed of random coil, 伪-helix and 尾-fold. According to the phylogenetic analysis of Erigeron breviscapus MYB and Arabidopsis MYB (AtMYB), it was found that the ebMYB gene was clustered with the subpopulations of AtMYB _ 4 and T _ MYB _ (11) ~ (121111) _ 2 in Arabidopsis thaliana. It was suggested that the cloned genes might share the same structure or function with these two groups. The results also showed that the constructed expression could be used for the efficient transformation of Erigeron breviscapus. Conclusion the MYB gene which may be involved in the regulation of phenylpropane metabolism or environmental response was cloned from Erigeron breviscapus for the first time.
【作者单位】：云南师范大学生命科学学院;生物能源持续开发利用教育部工程中心;
【基金】：国家自然科学基金资助项目(30660075,31360263) 云南省科技厅社会发展科技计划基础研究面上资助项目(2009CD052) 云南省基础研究计划资助项目(2011FA016)
【分类号】：S567.239

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