内源细胞分裂素调控油菜叶片衰老进程的研究

Delaying Leaf Senescence by Regulating Cytokinin Biosynthesis in Brassica napus

利用根癌农杆菌 (Agrobacteriumtumefaciens)介导法将嵌合基因PSAG1 2 ipt导入双低油菜 (Brassicanapus)品种H16 5 ,经双重PCR检测和基因组总DNASouthern杂交证实 ,共获得 11株转基因植株。生长观察结果显示 ,11株转基因植株生长发育正常 ,除 2株与对照没有明显的差异外 ,其余 9株叶片衰老时间明显延迟 ,主茎有柄叶衰老延迟 15～ 2 0d ,无柄薹叶延迟 2 0～ 30d ,长势较旺。对其中的 2株 (Line 5和Line 7)进行衰老生理指标测定的结果表明 ,转基因植株主茎基本定型叶上部叶片 ,叶绿素含量、细胞分裂素含量、SOD活性和MDA积累量与对照基本一致 ,转基因植株Line 5和Line 7基本定型叶下部第 5片叶叶绿素含量、细胞分裂素含量和SOD酶活性约为对照的 2～ 4倍、3～ 5倍和 3倍 ,MDA积累量仅为对照的 4 0 %和 70 %。

Leaf is the most important photosynthetic organ in rape and greatly contributes to the yield of seeds.So, the yield of seeds in rape is seriously influenced by the process of leaf senescence, which can be delayed by increasing endogenous or exogenous CTKs.By regulating CTKs biosynthesis, it is possible to get the leaf-senescence-delayed rape cultivar with higher yield of seeds.In this paper, some results were reported about rape transformation, and some developmental and physiological characteristics of the P SAG12 -ipt transgenic rapes.The chimeric gene P SAG12 -ipt was integrated into rape genome mediated by Agrobacterium tumefaciens EHA105 and 11 transgenic plants were successfully screened by double PCR and Southern blotting.The growth of these transgenic plants was observed in a systematic way.The results showed that the transgenic plants grew normally with more vigor than the controls and no morphological variation, the senescence delayed 15- 20 days for leaves with petiole in the main stem and 20-30 days for those without petiole.In different leaf-positions of 2 transgenic plants(Line 5 and Line 7), the contents of chlorophyll,cytokinins(ZRs and iPA),MDA and SOD activity were examined, which are the important physiological indexes about leaf senescence.The results showed that the contents of chlorophyll, cytokinins (ZRs and iPA) and SOD activity were obviously higher in the leaves below newly full-extended leaf than in the controls, while those in the extending leaves were identical to those in the controls.Whereas, the production of MDA was markedly inhibited.In the fifth leaf below newly full-extended leaf(Line 5 and Line 7), the contents of chlorophyll,cytokinins (ZRs and iPA),and SOD activity were about 2-4, 3-5 and 3 times of those in the controls respectively, while MDA production was only about 40% and 70% of the controls.These data prove that the ipt gene in the P SAG12 -ipt transgenic rapes can be controlled to express only in the senescent leaves and the leaf senescence can be physiologically delayed.