Preliminary Analysis Of A Leaf Senescence-associated Gene A4in Rice

Leaf is the place of photosynthesis and accumulation of photosynthate that is an veryimportant organs of plants. As the final stage of leaf development, leaf senescence playsimportant roles in the process of this period for plants. Leaf senescence can cause thedecline of photosynthesis, making it a limiting factor for crop yield. Rice is one of the mostimportant food crops in the world. Therefore, the study of leaf senescence mechanism isvery important to agricultural production.A4(LOC_Os12g43970) gene was selected from the GeneChip result of prematureover-expression transgenic line OX2. The expression of A4gene was increased14.5foldsin the GeneChip data. The GeneChip result of CREP (http://crep.ncpgr.cn/) suggested thatA4was a leaf senescence-associated gene.In the study of gene function, The expression effect of interference have morepersuasive than over-expression. Based on the previous results, we start by constructingRNAi construction and introducing it into the Zhonghua11by Agrobacterium-mediated.Detection of single copy and the expression of A4in flag leaves of14days after headingstage. And observe the leaf senescence of RNAi transgenic plants. Then construction of thelibrary and senescence yeast two-hybrid vector, screening the A4-interaction proteins. Itwas aimed to understand the mechanism of leaf senescence, and thus to produce thedelayed leaf senescence rice resources. The main results are as follows:1. Constructed the A4RNAi vector using pDS1301vector and introduced intoZhonghua11by EHA105Agrobacterium-mediated transformation, generated66transgenic plants,45of them are positive, the positive rate was68.18%;2. Six independent single-copy transgenic plants were confirmed by DIG-labeledSouthern blot analysis;3. The expression of A4was largely decreased in flag leaves of14days after headingstage in RNAi transgenic plants using quantitative real-time PCR of RNA (lowest reachedto25folds), which maked the interference was reliable. The expression of four single-copytransgenic plants has decreased to25~50%, suggesting that the RNAi effect was obvious.4. Constructed a yeast two-hybridAD-cDNA library using senescing leaves;5. Constructed the yeast transformation construction of A4full-length cDNA andtransformed into yeast, finding that the pGBKT7-A4protein has transactivation activitywithAH109.