Cloning and Expression Analysis of the Gene Encoding Flavonoid 3&#x02032;-Hydroxylase in Tea Plant (Camellia sinensis)

WANG Wenli; WU Zhijun; LIU Zhiwei; WANG Yongxin; LI Hui; CUI Xin; ZHUANG Jing

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Journal of Tea Science ›› 2017, Vol. 37 ›› Issue (1) : 108-118.

Cloning and Expression Analysis of the Gene Encoding Flavonoid 3′-Hydroxylase in Tea Plant (Camellia sinensis)

WANG Wenli, WU Zhijun, LIU Zhiwei, WANG Yongxin, LI Hui, CUI Xin, ZHUANG Jing^*

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Abstract

Flavonoid 3′-hydroxylase (F3′H) belongs to the family of cytochrome P450 monooxygenases (CYP450) and plays important roles in plant secondary metabolism and stress reponses. In this study, a gene encoding F3’H-like protein was cloned by RT-PCR method using a cDNA template from tea (Camellia sinensis) cultivar ‘Longjing43’. This gene is named as CsF3′H1. Sequence analysis showed that the open reading frame of CsF3’H1 was 1,530 bp length, encoding 509 amino acids. CsF3′H1 protein contains the conserved binding domain of P450 enzyme. Analysis of phylogenetic tree showed that CsF3'H1 protein has high similarity with CsF3'H3 protein and has low similarity with CsF3'H2 protein. Multiple alignments showed that CsF3'H1 protein has high similarity with the F3′H homologs from Sorghum bicolor, Actinidia chinensis, Populus trichocarpa, Arabidopsis thaliana, and Vitis vinifera (66.48% identity). CsF3'H1 protein contains the characterized motifs of F3'H-type protein. Analysis of amino acid composition, physical and chemical properties, hydrophilicity/hydrophobicity, and disordered residues of CsF3′H1 protein showed that the disordered residues of CsF3′H1 protein are not obvious and most amino acids of CsF3′H1 protein are hydrophilic. The expression profiles of CsF3′H1 gene in different tissues of tea plant or under hormonal treatments were detected using quantitative real-time PCR analysis. Results showed that: CsF3'H1 gene has the highest expression level in the first leaf. The tissue expression profiles showed that the successive order of CsF3'H1 gene expression levels was the first leaf > the second leaf > the third leaf > the fourth leaf > stem > roo > old leaf. The expression level of CsF3'H1 gene was highest under SA treatment, which was 2.24 times high than the control. CsF3'H1 gene had the lowest expression level under MeJA treatment.

Key words

flavonoid 3′-hydroxylase / hormonal treatments / phylogenetic tree / tissue expression / Camellia sinensis

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WANG Wenli, WU Zhijun, LIU Zhiwei, WANG Yongxin, LI Hui, CUI Xin, ZHUANG Jing. Cloning and Expression Analysis of the Gene Encoding Flavonoid 3′-Hydroxylase in Tea Plant (Camellia sinensis)[J]. Journal of Tea Science. 2017, 37(1): 108-118

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