中国野生毛葡萄‘丹凤-2’3个芪合酶基因表达与功能分析

Expression and Function Analysis of Three Stilbene Synthase Genes from Vitis quinquangularis Rehd.Accession‘Danfeng-2'

为了研究中国野生毛葡萄(Vitis quinquangularis Rehd.)‘丹凤-2’3个芪合酶基因的表达与功能,该研究采用同源克隆技术分离了毛葡萄‘丹凤-2’3个芪合酶基因VqSTS21、VqSTS30和VqSTS32(GenBank登录号分别为JQ868677、JQ868668和JQ868666),3个基因的cDNA全长均为1 179bp,编码392个氨基酸。以抗葡萄白粉病(Uncinula necator)的毛葡萄‘丹凤-2’和不抗病的欧洲葡萄‘赤霞珠’为材料分别进行接种白粉菌和ABA、SA、MeJA、高温、低温和高盐胁迫处理,利用实时定量PCR检测这3个基因在胁迫处理下的表达情况;同时利用农杆菌介导法将这3个基因分别转入模式植物烟草中,检测这3个基因在烟草中的表达产物,分析比较这3个基因的表达功能。结果显示:在2个供试材料中,葡萄白粉菌胁迫下芪合成酶VqSTS32诱导表达量高于VqSTS21和VqSTS30;在非生物胁迫处理下,芪合酶基因VqSTS32对高温处理反应最敏感。采用高效液相色谱分析检测转3个基因烟草的结果显示,转VqSTS32基因烟草比转VqSTS30基因烟草植株中白藜芦醇的累积量高。研究表明,3个基因中VqSTS32具有较高的抗葡萄白粉菌特性并能在转基因烟草中表达出较高的反式云杉新苷产物,这为进一步利用VqSTS32转目的植物葡萄基因研究提供了依据。

The purpose of this paper is to study the expression and function of VqSTS21,VqSTS30 and VqSTS32genes,which cloned from powdery mildew-resistant Chinese wild Vitisquinquangularisaccession‘Danfeng-2’.Sequence analysis indicated that their cDNA has equal length of 1 179 bp and encodes392 amino acid residues（GenBank accession No.JQ868677,JQ868668 and JQ868666）.qRT-PCR showed the expression patterns of VqSTS21,VqSTS30 and VqSTS32in‘Danfeng-2’and powdery mildew-susceptible Cabernet Sauvignon（Vitis vinifera L.）under biotic stress（Uncinula necator）and abiotic stress（ABA,SA,MeJA,heat,low temperature and NaCl）.HPLC was employed to analysis resveratrol contents of these three genes overexpressed in transgenic tobacco plants.The results showed that the expression of VqSTS32 was higher than that of other two genes in two grape germplasms tested after treated with biotic stress and heat.Compared with the VqSTS21 and VqSTS30-overexpressed lines,trans-resveratrol was thehighest accumulation in the VqSTS32-overexpressed lines of transgenic tobacco plants.Our results suggested that VqSTS32 gene was involved in the resveratrol synthesis pathway in grapes,and played an important role in response to U.necator.