Cloning of CYP71 gene in Malania oleifera and expression analysis in different developmental periods

Abstract

Amino acids-derived cyanogenic glucosides catalyzing by plant cytochrome P450 enzymes are plant secondary metabolism,which is related to plant defense and anti-stress.A gene of cytochrome P450(namely MoCYP71,GenBank ID is MK172858) from Malania oleifera was isolated and cloned through analyzing the transcriptome of M.oleifera,then its function was analyzed by bioinformatics analysis,and the expression of MoCYP71 in different developmental periods of fruit was detected.The results were as follows: MoCYP71 gene has 1 572 bp,encodes 523 amid acids,and its sequence of cDNA has 88% identity with the CYP71 cDNA of Coffea eugenioides(XM027319282) and Coffea arabica(XM027213456);The relative molecular weight of MoCYP71 protein was 58 976.54,the theoretical pI was 8.10,the molecular formula was C(2675)H4184N704O744S27,the instability index(Ⅱ) was 40.84,and the protein belonged to an instable protein;There had no siginal peptide in this protein sequence,and the protein exists in the secretion pathway and contained two transmembrane structure,located in the protein sequences of 20-37 and 311-333,and the transmembrane sites anchored in the organelle membrane;MoCYP71 had all the conserved domain of CYP family,containing prolinerich region(PPSPPRLP),K helix region(KETFR),I helix region(GGIDTS),PERF domain(PERF) and the main identified feature heme-binding region(FGAGRRICPG),and the protein MoCYP71 and the CYP71 E protein in Theobroma cacao,Durio zibethinus,Sorghum bicolor(GenBank ID is EOX92908.1,XP022773875.1 and AAC39318.1,respectively) clustered into one clan;The expression level of MoCYP71 gene reduced gradually in the first three months after blossom fading,particularly,1st month the fruit after blossom fading> 2nd month> 3rd month,however,its expression level increased sharply in 4th month.The present study had a important significance to defense to insect pests,tissue ripening and secondary metabolism digging in Malania oleifera.