调控转基因植物EβF释放量的抗蚜虫研究

(E)-β-farnesene (EβF), as the main or generally only component of the aphid alarm pheromones, can cause repellence of aphids and also function as a kairomone in attracting aphid predators. Transgenic plant engineered with EβF synthase genes is an efficient strategy for aphid management, in which EβF synthase enzymes convert cytosolic farnesyl diphosphate (FPP) to the acyclic sesquiterpene EβF. However, relatively low EβF emission level of transgenic plants was observed. The availability of FPP supply in the cytoplasm, as well as the absence and/or relatively low catalyzing activity of the plastid isoform of FPP synthase have become the major limiting factors in biosynthesis of EβF in plants for aphid control. In our previous studies, the FPP synthase gene Atfps1 and EβF synthase gene MaβFS1 have been isolated from Arabidopsis plants and peppermint, respectively; and the chloroplast-targeting property of a chloroplast transit peptide (CTP) from wheat small subunit of Rubisco (rbcS) has been identified.Basing on these research foundations, to obtain the best strategy for transgenic plants emitting sufficient EβF against aphid infestation, the strategies of overexpressing MaβFS1 and/or both of Atfps1 and MaβFS1 in tobacco cytoplasm or chloroplast will be taken into account in this project, respectively. Then, the EβF emission levels and aphid/parasitoid bioassays of these transgenic lines will be comprehensively investigated. This research will lay the theoretical foundation for effectively engineering with EβF synthase gene against aphid infestation in other economically important crop plants.

蚜虫是重要的农业害虫，EβF是蚜虫报警信息素的主要成份，可趋避蚜虫并吸引天敌。植物中表达EβF合成酶基因以催化FPP合成EβF是控制蚜虫危害的重要策略，但仍存在两个问题：一是细胞质FPP供应量不足；二是质体尽管有足够的前体合成FPP，但FPP合成酶缺失或者催化效率低，导致植物EβF释放量偏低。申请者前期分离了薄荷EβF合成酶基因MaβFS1、拟南芥FPP合成酶基因Tafps1及水稻rbc S启动子，并从小麦分离到可以介导蛋白定位于叶绿体的转导肽CTP。在此基础上，本项目拟分别在烟草细胞质及叶绿体中表达单价基因MaβFS1、双价基因Tafps1和MaβFS1、以及使用不同启动子获得不同的转基因植株，通过系统比较不同转基因植株中EβF及其它萜类化合物的释放量及其与抗蚜性水平的关系，进而筛选出提高植物EβF释放量控制蚜虫危害的最佳策略。此项目为EβF合成酶基因在其它作物中的高效抗蚜育种提供依据。

EβF合成酶可催化植物细胞质中的FPP合成EβF，植物中表达该基因可减轻蚜虫危害。研究表明受体植物细胞质中FPP供应量不足造成EβF释放量偏低，影响蚜虫防治效果。在前期研究的基础上，本项目：1）利用CaMV 35S启动子，创制了细胞质及叶绿体中表达单价基因MaβFS1、双价基因AtFPS1和MaβFS1的烟草植株；2）利用水稻rbc S启动子，创制了细胞质及叶绿体中表达单价基因MaβFS1、双价基因AtFPS1和MaβFS1的烟草植株；3) 通过GC-MS分析和抗蚜鉴定，系统比较了不同类型转基因植株中EβF释放量及抗蚜水平。结果表明，水稻rbcS基因启动子可驱动EβF合成酶基因在烟草绿色组织中特异表达，与CaMV 35S的同类型转基因植株相比，启动子对转基因植株的EβF释放量没有明显影响。叶绿体中同时表达AtFPS1和MaβFS1，利用AtFPS1催化叶绿体内的IPP和DMAPP合成FPP，可显著提高烟草幼苗的EβF释放量。叶绿体中表达双价基因的烟草幼苗可以趋避蚜虫，而其它类型的烟草植株驱蚜效果不显著。本项目的研究结果为EβF合成酶基因在其它农作物中的转基因抗蚜育种提供了策略，即将EβF的代谢途径从细胞质转移至质体，在质体中同时表达EβF和FPP合成酶基因。