Bioinformatical and Expression Analysis of Flavonoid-3'5'-Hydroxylase Gene from Pigmented Potato Named ‘ZhuanXinWu’
Flavonoid-3'5'-hydroxylase (F3'5'H) is the key enzyme which can synthesize delphinidin-3-glucoside in anthocyanin biosynthesis.In this study,the complete cDNA of F3'5'H was isolated from the tuber of pigmented potato named ‘ZhuanXinWu’ (Solanum tuberosum L.) by reverse transcription PCR (RT-PCR).The F3'5'H sequence information and expression pattern were analyzed by tools of bioinformatics and Real time Quantitative PCR (qRT-PCR),to find out the role of F3'5'H gene in the pigmented potato anthocyanins synthesis and the expression pattern.The complete F3'5'H cDNA had 1 720 base pair and encoded an poly peptide composed of 509 amino acid residues.Through multiple sequences alignment of F3'5'H,a homology tree was constructed.The F3'5'H of ‘ZhuanXinWu’ were clustered together with Solanaceae plants and had the closest phylogenetic relationship,next was other dicotyledon.According to sequence analysis,F3'5'H had signal peptide and obvious transmembrane structure region.It belonged to stable hydrophilic secretory protein and probably located in cytoplasm.These results illuminated that F3'5'H was transferred to other cell organelles and exerted its function after precursor was synthesized in rough endoplasmic reticulum of cytoplasm.α-helix and random coil were primary secondary structural components of F3'5'H.The F3'5'H protein had the highly conserved sites of ‘PPGP’,‘AGTDT’ and ‘FGAGRRICAG’ and had only one functional domain.The functional domain of F3'5'H matched the functional construction of Cytochrome P450,therefore F3'5'H protein belonged to Cytochrome P450 superfamily.The mRNA expression analysis by real-time quantitative PCR demonstrated that the accumulation of F3'5'H transcripts in tuber were much more than that in leaf and stem.It indicated that the expression ofF3'5'H gene was followed by anthocyanin accumulation.The activity of F3'5'H was dramatically and positively correlated with the content of anthocyanin.
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