Cloning of MYB gene and construction of greenfluorescent protein expression vector in Erigeron breviscapus / 中草药

ABSTRACT

Objective To study the role of MYB transcription factor in phenylpropane biosynthesis pathway and the response to biotic or abiotic stress, a full length sequence of putative MYB gene was cloned in Erigeron breviscapus. Methods Based on a partial sequence of putative MYB gene from transcriptome we have previous reported, the full length cDNA was cloned by RACE method in E. breviscapus. According to the sequence of cloning cDNA, the nucleotide sequence similarity, physicochemical properties, hydrophobicity, transmembrane structure, secondary structure and tertiary structure were predicted and analyzed by various softwares. Meanwhile, multiple sequence alignment of cloned MYB gene was performed, and the phylogenetic tree was constructed. In addition, the fusion expression vector of this gene and green fluorescent protein was constructed. Results The MYB gene was cloned and named eBMYB06. The open reading frame was 783 bp, encoding 260 amino acid residues with a relative molecular mass of 63.80 kDa and a theoretical pI of 5.18 which proved to be a stable protein. The secondary structure of the protein is mainly composed of irregular coil, alpha helix and beta fold. According to the result of phylogenetic tree alignment of E. breviscapus with R2R3MYBs from Arabidopsis thaliana, the cloned MYB gene was clustered to two subsets of R2R3-MYB genes from A. thaliana, which suggested that the cloned MYB gene would be similar with the two groups in structure and function, which would be involved in response to biotic and abiotic stress or phenylpropane biosynthesis pathway, respectively. Further experimental results show that the constructed expression can be used for the efficient transformation in E. breviscapus. Conclusion For the first time, a MYB gene was cloned may be involved in the phenylpropanoid metabolism or gene metabolism or gene regulation in response to the environment in E. breviscapus.