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首页> 外文期刊>Plant Cell Reports >Generation of selectable marker-free sheath blight resistant transgenic rice plants by efficient co-transformation of a cointegrate vector T-DNA and a binary vector T-DNA in one Agrobacterium tumefaciens strain
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Generation of selectable marker-free sheath blight resistant transgenic rice plants by efficient co-transformation of a cointegrate vector T-DNA and a binary vector T-DNA in one Agrobacterium tumefaciens strain

机译：通过在一株根癌土壤杆菌菌株中共整合载体T-DNA和二元载体T-DNA的有效共转化，产生无选择标记的抗白叶枯病抗性转基因水稻植株

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摘要

Co-transformation of Oryza sativa L. var. Pusa Basmati1 was done using an Agrobacterium tumefaciens strain harbouring a single-copy cointegrate vector and a multi-copy binary vector in the same cell. The T-DNA of the cointegrate vector pGV2260::pSSJ1 carried the hygromycin phosphotransferase (hph) and β-glucuronidase (gus) genes. The binary vector pCam-chi11, without a plant selectable marker gene, harboured the rice chitinase (chi11) gene under maize ubiquitin promoter. Co-transformation of the gene of interest (chi11) with the selectable marker gene (hph) occurred in 4 out of 20 T0 plants (20%). Segregation of hph from chi11 was accomplished in two (CoT6 and CoT23) of the four co-transformed plants in the T1 generation. The selectable marker-free (SMF) lines CoT6 and CoT23 harboured single copies of chi11. Homozygous SMF T2 plants were established in the lines CoT6 and CoT23. Northern and Western blot analysis of the homozygous SMF lines showed high level of transgene expression. In comparison to untransformed controls, chitinase specific activity was 66- and 22-fold higher in the homozygous SMF T2 plants of lines CoT6 and CoT23, respectively. The lines CoT6 and CoT23 exhibited 38 and 40% reduction in sheath blight disease, respectively.
机译：水稻的共转化Pusa Basmati1使用根癌农杆菌菌株完成，该菌株在同一细胞中具有单拷贝共整合载体和多拷贝二元载体。共整合载体pGV2260 :: pSSJ1的T-DNA带有潮霉素磷酸转移酶（hph）和β-葡萄糖醛酸苷酶（gus）基因。没有植物选择标记基因的二元载体pCam-chi11在玉米遍在蛋白启动子下带有水稻几丁质酶（chi11）基因。在20株T0 植物中，有4株（20％）发生了目的基因（chi11）与选择标记基因（hph）的共转化。在T1世代的4种共转化植物中，有2种（CoT6和CoT23）完成了chi11中hph的分离。可选的无标记（SMF）行CoT6和CoT23包含chi11的单个副本。在CoT6和CoT23品系中建立了纯合SMF T2 植物。纯合SMF系的Northern和Western印迹分析显示高水平的转基因表达。与未转化的对照相比，在品系CoT6和CoT23的纯合SMF T2 植物中，几丁质酶的比活性分别高66和22倍。 CoT6和CoT23品系的鞘枯萎病分别减少了38％和40％。

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