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期 刊 ：植物遗传资源学报 2014年 15卷 3期

关键词：青稞(裸大麦)；C4H基因；同源克隆；RACE；实时荧光定量PCR；胚乳发育期；

Keywords:Qingke(hulless barley), cinnamate-4-hydroxylase(C4H) gene, Homologous cloning, Rrapid amplification of cDNA end(RACE), Real-time-fluorescence quantitative PCR, Endosperm development,

摘 要 ：为揭示大麦中黄酮合成的分子调控机制，利用反转录PCR结合同源克隆和RACE技术首次从青稞（裸大麦）叶片中克隆得到肉桂酸-4-羟化酶基因(HvC4H )的全长cDNA序列（Genbank登录号: KF927086），总长度1951bp，ORF为1518bp，编码505个苯丙氨酸代谢氨基酸，等电点PI=9.01，平均亲水指数(GRAVY)为-0.170，属于亲水性碱性蛋白，高级结构分析其具有细胞色素P450家族保守域及C4H特异的功能性活性位点。利用实时荧光定量PCR分析胚乳发育5个时期不同组织(茎、叶及籽粒)的表达情况，结果显示HvC4H基因在青稞胚乳发育期的表达情况存在着明显的组织差异性，HvC4H在胚乳发育期茎中的表达占优势，为[删除]通过控制大麦中C4H基因的表达从而提高大麦黄酮的含量奠定分子基础，对于改良大麦的品质、抗性、生长发育等性状具有重要意义。

Abstract:In order to reveal the molecular regulatory mechanisms of flavonoids biosynthesis in barley(Hordeum vulgare), the full length cDNA encoding of cinnamate-4-hydroxylase gene(HvC4H) was cloned from leaves of Qingke(hulless barley) through the methods of RT-PCR combining homologous clone and RACE technologies(Genbank accession number: KF927086). The full length was 1951bp , open reading frame (ORF) was 1518bp encoding 505 amino acids. Isoelectric point was 9.01 and grand average of hydropathicity (GRAVY) was -0.170, it meant the HvC4H was hydrophilic and alkaline protein. Advanced structure analysis showed HvC4H gene including family CYP450 conserved domains and specific functional active sites. The expression of HvC4H gene was analyzed in different tissues(stem, leaf and kernel) during 5 periods in endosperm development by real-time-fluorescence quantitative PCR(RTF-qPCR), the results showed obvious differences and the expression in stem was predominant . The study could lay the molecular foundation to raise the content of flavonoid in barley by controlling the expression of HvC4H gene and it provided useful informations to improve the quality, resistibility and growth of barley.

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