Cloning and expression pattern analysis of chalcone synthase gene in Microcos paniculata / 中草药

ABSTRACT

Objective To clone the full-length cDNA of MpCHS and analyze its expression pattern in different parts of Microcos paniculata and different growth periods of M. paniculata leaves. Methods Based on the transcriptome data of M. paniculata, we designed specific primers for MpCHS gene. The full-length cDNA of MpCHS was amplified by PCR and the positive clones were then sequenced, analyzed, and constructed prokaryotic expression vector. The bioinformatics analysis of MpCHS was also performed. Meanwhile, the mRNA expression of MpCHS was detected using real-time quantitative PCR. Results The relative molecular mass was 42 700, and its theoretical isoelectric point was 6.11, with three conserved functional active sites (165 C, 304 H, and 337 N) of the CHS family proteins and the tag sequence of RLMMYQQGCFAGGTVLR and GVLFGFGPGL. Phylogenetic tree analysis showed that MpCHS had close relationship with woody plants such as cocoa and upland cotton. We successfully cloned the full-length cDNA of MpCHS (GenBank KY472608). It had an ORF of 1 176 bp which encoded a protein of 391 amino acid residues. RT-qPCR results showed that MpCHS was expressed in all parts of M. paniculata and its expression in leaves was gradually decreased along with its development. Conclusion MpCHS is cloned from M. paniculate for the first time, and the gene expression pattern of MpCHS in different parts of M. paniculata and different growth periods of M. paniculata leaves was analyzed. This study facilitates the further purification and functional validation of MpCHS protein and provides reference for further analysis of flavonoids biosynthesis pathway in M. paniculata.