RNAi沉默Fa4CL基因对草莓果实花色苷代谢的影响

Influences of RNAi-induced Fa4CL silencing on anthocyanin metabolism in strawberry fruit

【目的】探讨Fa4CL与花色苷代谢之间的关系,构建RNA干扰载体转染草莓果实,研究Fa4CL基因在草莓花色苷代谢中的功能。【方法】用RT-PCR方法从‘阿尔比’草莓(Fragaria×ananassa‘Albion’)果实中克隆Fa4CL基因。构建不含内含子的发夹结构干扰载体p BI-4CLi,将其转入农杆菌GV3101,采用无菌注射器注入到授粉后14 d的‘阿尔比’草莓果实中转染,与对照果比较表型变化、物质变化,半定量RT-PCR检测Fa4CL基因的表达量的变化。【结果】克隆的Fa4CL的CDS,长1638 bp,编码545个氨基酸。与森林草莓Fv4CL2基因序列的同源性较高,为98.4%。p BI-4CLi转染果与对照果相比,转染果的果皮红色变浅。进一步利用RT-PCR分析发现,p BI-4CLi转染果Fa4CL基因表达量明显低于对照果。分析果实花色苷含量发现,p BI-4CLi转染果的花青素糖苷和花葵素糖苷分别比对照果降低了93.5%和97.0%,差异均达到显著水平。【结论】RNA干扰载体转染草莓果实的结果证明Fa4CL成功沉默,Fa4CL基因与花色苷的合成关系密切。

【Objective】We studied the role that Fa4 CL gene plays in the anthocyanin metabolism of straw-berry by constructing the interference vector that infected the fruit. 【Methods】The Fa4 CL gene wascloned from‘Albi'strawberry fruit(Fragaria × ananassa‘Albion') using RT-PCR. We built intron-freehairpin interference vector p BI-4CLi,transformed it into agrobacterium GV3101,injected the bacteriuminto strawberry fruit with a sterile syringe 14 days after pollination,and then investigated the changes inphenotypes,materials,and 4CL gene expression in the transfected fruit and control. 【Results】Thecloned Fa4 CL c DNA has 1638 bp and encodes 545 amino acids. Its sequence is more closely related toFragaria vesca,with homology up to 98.4%. The red color of transfected fruit peels became shallow. Fur-ther analysis with RT-PCR indicated that the expression of Fa4 CL gene was significantly depressed inthe transfected fruit. The content of anthocyanin glucoside and pelargonidin glycosides decreased by93.5% and 97.0% in p BI-4CLi transfected fruit compared with that in the control fruit at the significantlevel.【Conclusion】The experimental results show that the Fa4 CL gene was successfully silenced and it isclosely associated with the synthesis of anthocyanins in strawberry fruit.