Cloning and Expression Analysis of Transcription Factor MYB4 Gene from Angelica sinensis

YANG Xue; LUO Jun; YANG Caixia; WANG Zhenheng; XIA Qi; WANG Yinquan

doi:10.15933/j.cnki.1004-3268.2018.12.008

Abstract：To explore the structure and function of MYB protein involved in the biosynthesis of ferulic acid in Angelica sinensis, the transcription factor gene of MYB4 was rapidly isolated and cloned from Angelica sinensis, and sequence analysis and expression analysis of MYB4 gene were performed.According to the DNA conserved domain of MYB4 transcription factor, combined with the principle of homologous cloning and RACE, the full-length cDNA of MYB4 gene was cloned from Angelica sinensis by RT-PCR;The basic properties, structural characteristics and biological functions of the protein encoded by MYB4 gene were preliminarily predicted by using bioinformatics analysis tools;The expression characteristics of MYB4gene were analyzed by qRT-PCR;Recombination technology was used to assemble prokaryotic expression vector pGEX-4T-3-As MYB4, which was transformed into Escherichia coli BL21(DE3) by heat shock, and IPTG was used to induce the expression of recombinant protein.The results showed that a MYB4 gene was cloned and named As MYB4, which was registered in GenBank and the accession number was MG736315;Sequence analysis showed that the full-length cDNA of As MYB4 gene was 1 189 bp, including a 804 bp open reading frame, and encoding 267 amino acids.It was a typical member of R2R3-MYB transcription factor family;The homology between As MYB4 and Daucus carota MYB308 reached 93％, and As MYB4had the closest relationship with Daucus carota MYB308.Tissue specific expression analysis showed that transcription level of As MYB4 gene was highest in leaves of Angelica sinensis, which was 1.1 and 2.0times of petioles and roots, respectively;Prokaryotic expression analysis showed that As MYB4 protein existed as inclusion bodies with a molecular mass of 30 ku.