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以‘清榄一号’橄榄(Canarium album)果肉为材料,通过RACE技术克隆得到橄榄类黄酮3’-羟化酶(F3’H)cDNA序列全长,命名为CaF3’H(GenBank登录号KY189088.1)。CaF3’H全长为1 649 bp,包含一个1 554 bp开放阅读框(ORF),编码517个氨基酸,5’和3’非编码区(UTR)长度分别为40 bp和55 bp,编码的氨基酸序列含有P450结构域和半胱氨酸亚铁血红素结合域保守区。氨基酸多重比较分析表明:橄榄和芒果的氨基酸序列相似性为78.89%,较之于其他物种相对最高。蛋白结构预测结果显示,CaF3’H蛋白呈弱亲水性,存在跨膜结构域,没有信号肽。实时荧光定量PCR分析显示,CaF3’H在‘清榄一号’和‘檀香’2个橄榄栽培品种的表达量在花后50 d达到最大值,此后其表达量随果实的成熟呈现出波动,总体表达量比花后50 d都低,这与橄榄生长发育时期多酚含量的变化趋势相符,推测CaF3’H基因在调控果实多酚含量过程中发挥着重要作用。
The full length of the cDNA sequence of the olive flavonoid 3’-hydroxylase (F3’H) was cloned by RACE using Cauliflower ’Canarium album’ pulp as CaF3’H (GenBank accession number KY189088 .1). CaF3’H was 1 649 bp in length and contained a 1 554 bp open reading frame (ORF) encoding 517 amino acids. The 5 ’and 3’ non-coding regions (UTRs) were 40 bp and 55 bp respectively. The encoded amino acids The sequence contains the P450 domain and the conserved region of the cysteine heme binding domain. Multiple amino acid analysis showed that the amino acid sequence similarity between olive and mango was 78.89%, which was the highest relative to other species. The results of protein structure prediction showed that CaF3’H protein was weakly hydrophilic and had a transmembrane domain and no signal peptide. Real-time PCR analysis showed that the expression level of CaF3’H in two olive cultivars of ’Qinglou No.1’ and ’Sandalwood’ reached the maximum at 50 days after flowering, and the expression level of CaF3’H fluctuated with the ripening of fruits , The overall expression level was lower than that at 50 d after anthesis, which was consistent with the trend of polyphenol content during the growth and development of olives. It is speculated that CaF3’H gene plays an important role in the regulation of polyphenol content in fruit.