HvABA8′OH-1基因RNAi载体的构建及遗传转化

Construction of RNAi Vector Targeting HvABA8'OH-1 and Its Genetic Transformation

穗发芽严重影响作物的产量和品质,种子休眠性弱是穗发芽形成的主要原因之一.ABA是维持胚休眠抑制萌发的重要激素,大麦HvABA8′OH-1基因编码控制内源ABA主要分解代谢途径的关键酶.以双元载体pGreenⅡ为骨架载体,分别选取HvABA8′OH-1的保守区序列(293bp)和特异区序列(474bp),水稻胚乳特异性启动子pGlub,成功构建了种子特异性RNA干扰表达载体pOHC和pOHS.利用农杆菌介导法将载体转入水稻,获得植株50株.经鉴定,其中18株为转基因阳性植株.研究结果为进一步分析HvABA8′OH-1基因的生物学功能,以及通过基因工程途径建立作物抗穗发芽育种新方法提供了理论依据和材料.

Pre-harvest sprouting（PHS） seriously affects the quality and yield of cereal crops.The weak dormancy of seed is one of the major reasons causing PHS.As an important phytohormone,abscisic acid（ABA） plays an important role in seed development which is closely involved in the onset and maintenance of dormancy,repressing germination.HvABA8＇OH-1encoded ABA 8＇-hydroxylationen is the key gene controlling ABA catabolism in barley.In this study,two RNA interference（RNAi） expression vectors targeting the conservative and specific regions of HvABA8＇OH-1 were constructed,respectively.The vectors,namely,pOHC and pOHS,were composed of inversely-repeated sequence fragments,an endosperm specific promoter pGlub and a hygromycin resistance gene as a selectable marker.The RNAi expression vectors were transformed into rice by Agrobacterium-mediated transformation.50 plants were obtained and 18 of them were identified as transgenic plants by PCR.The above results might be useful for the further study of biological function of HvABA8＇OH-1 and establishment of a new way for controlling PHS by genetic engineering.