Molecular cloning and sequence analysis of(S)-N-methylcoclaurine-3'-hydroxylase gene in Coptis chinensis / 中草药

ABSTRACT

Objective To clone and sequence the cDNA encoding(S)-N-methylcoclaurine-3'-hydroxylase from Coptis chinensis.Methods The cDNA,encoding(S)-N-methylcoclaurine-3'-hydroxylase,was amplified by RT-PCR with cDNA library of tender leaf as the template.Results The full-length cDNA of(S)-N-methylcoclaurine-3'-hydroxylase(named as CYP80B3) had 1 680 bp with an open reading frame encoding 488 amino acids of protein.The CYP80B3 had 95%,82%,70%,and 68% amino acid sequence homology to the sequence of(S)-N-methylcoclaurine-3'-hydroxylase from C.japonica,Thalictrum flavum,Eschscholzia californica and Papaver somniferum,respectively.The sequence was reported to the GenBank and coded as EF492879.Comparison of sequence with(S)-N-methylcoclaurine-3'-hydroxylase from C.japonica showed CYP80B3 possessed the same functional regions involved with 3'-hydroxylation of(S)-N-methylcoclaurine.Conclusion The cDNA encoding CYP80B3 from C.chinensis was cloned and reported.This work underlays the first step for exploring the pathway of benzylisoquinoline alkaloid biosynthesis and for improving the content of benzylisoquinoline alkaloid in C.chinensis.