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园艺学报 ›› 2015, Vol. 42 ›› Issue (8): 1495-1504.doi: 10.16420/j.issn.0513-353x.2015-0177

• 蔬菜 • 上一篇    下一篇

茄子SmMsrA基因VIGS表达载体的构建及表达分析

赵祯,刘富中,张映,齐东霞,陈钰辉,连勇   

  1. 中国农业科学院蔬菜花卉研究所,北京 100081
  • 出版日期:2015-08-25 发布日期:2015-08-25
  • 基金资助:
    国家‘863’计划项目(2012AA100202);中国农业科学院科技创新工程项目(CAAS-ASTIP-IVFCAAS);科技部国际合作项目(2011DFR31180);农业部园艺作物生物学与种质创制重点实验室项目

VIGS Expression Vector Construction and Expression Analyses of SmMsrA Gene in Eggplant

ZHAO Zhen, LIU Fu-Zhong, ZHANG Ying, QI Dong-Xia, CHEN Yu-Hui, LIAN Yong   

  1. Institute of Vegetables and Flowers,Chinese Academy of Agricultural Sciences,Beijing 100081,China
  • Online:2015-08-25 Published:2015-08-25

摘要: 利用RT-PCR从茄子单性结实品系D-10中获得甲硫氨酸亚砜还原酶A基因(SmMsrA)的编码区。通过Gateway同源重组技术,构建了3个分别含SmMsrA基因不同片段的烟草脆裂病毒(Tobacco rattle virus,TRV)介导的基因沉默(VIGS)表达载体pTRV2-SmMsrAi。表达载体转入农杆菌GV3101后,用注射压迫法侵染茄子叶片。采用表型观察、TRV病毒分子检测和qRT-PCR技术,评价构建的VIGS体系对SmMsrA基因的沉默效果。结果表明,报告基因PDS沉默后叶片呈现明显的光漂白现象,SmMsrA基因沉默后叶片呈花叶状,果实变小,叶片和果实中的SmMsrA基因的表达量明显降低,表明SmMsrA基因是正向调控茄子果实发育的相关基因。

关键词: 茄子, SmMsrA基因, VIGS, Gateway技术, 果实发育

Abstract: The coding sequence of methionine sulfoxide reductase A(SmMsrA)was amplified from eggplant(Solanum melongena L.)parthenocarpic line D-10 using reverse transcription polymerase chain reaction(RT-PCR)method. Through the Gateway recombination technology,three TRV(Tobacco rattle virus)-mediated VIGS expression vectors were constructed,pTRV2-SmMsrAi,containing different fragments of SmMsrA gene. The VIGS expression vectors were transformed into Agrobacterium strain GV3101,and then inoculated eggplant leaves by Agro-syringe. Phenotypes observation,TRV molecular detection and qRT-PCR thechnology were used to observe and evaluate the VIGS system for silencing the SmMsrA gene effection. The results showed that the positive control of silencing reporter gene PDS showed photo-bleaching phenotype in eggplant leaves;Silencing SmMsrA gene were observed pale yellow(mosaic)leaves and bore smaller fruits;The expression of SmMsrA gene was decreased obviously in the leaves and fruits of the infected eggplant plants. These results suggested that theSmMsrA gene was a related gene in positive regulation of fruit development in eggplant.

Key words: eggplant, SmMsrA gene, VIGS, Gateway technology, fruit development

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