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Enhanced Agrobacterium-mediated Transformation of Embryogenic Calli of Upland Cotton via Efficient Selection and Timely Subculture of Somatic Embryos

文献类型： 外文期刊

作者： Wu, Shen-Jie 1 ; Wang, Hai-Hai 1 ; Li, Fei-Fei 1 ; Chen, Tian-Zi 1 ; Zhang, Jie 1 ; Jiang, Yan-Jie 1 ; Ding, Yezhang 1 ;

作者机构： 1.Nanjing Agr Univ, Natl Key Lab Crop Genet & Germplasm Enhancement, Nanjing 210095, Peoples R China

2.Shanxi Acad Agr Sci, Cotton Res Inst, Yuncheng 044000, Peoples R China

关键词： plant regeneration;transformation efficiency;Embryogenic Calli;selection cultivation;embryogenic calli morphology

期刊名称：PLANT MOLECULAR BIOLOGY REPORTER （ 影响因子：1.595； 五年影响因子：2.042 ）

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收录情况： SCI

摘要： Agrobacterium-tumefaciens-mediated transformation of cotton embryogenic calli (EC) was enhanced by choosing appropriate EC and improving efficiency of coculture, selection cultivation, and plant regeneration. After 48-h cocultivation, the number of beta-glucuronidase (GUS)-positive calli characterized by yellow, loose, and fine-grained EC was twofold greater than that of gray, brown, and coarse-granule EC. It indicated that efficiency of transient transformation was affected by EC morphology. And transient transformation efficiency was also improved by cocultivation on the medium adding 50 mg l(-1) acetosyringone at 19 C for 48 h. Subculturing EC on the selection medium with low cell density was beneficial to production of more kanamycin-resistant (Km-R) calli lines. From an original 0.3-g EC, an average of 20 Km-R calli lines were obtained from a selection dish and the GUS-positive rate of Km-R clones was 81.97%. A large number of normal plants were rapidly regenerated on the differentiation medium with dehydration treatments and the GUS-positive rate of regeneration plants was about 72.60%. Polymerase chain reaction analysis of GUS-positive plantlets revealed a 100% positive detection rate for neomycin phosphotransferase II gene and uidA. Southern blot of transgenic plants regenerated from different Km-R calli lines demonstrated that the target gene, mostly with the low copy number, has been integrated into the cotton genome.

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