1. Enhanced Agrobacterium-mediated Transformation of Embryogenic Calli of Upland Cotton via Efficient Selection and Timely Subculture of Somatic Embryos.
- Author
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Shen-Jie Wu, Hai-Hai Wang, Fei-Fei Li, Tian-Zi Chen, Jie Zhang, Yan-Jie Jiang, Yezhang Ding, Wang-Zhen Guo, and Tian-Zhen Zhang
- Subjects
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AGROBACTERIUM , *COTTON , *SOMATIC embryogenesis , *PLANT cell culture , *RHIZOBIACEAE - Abstract
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 Ī²-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 lā1 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. [ABSTRACT FROM AUTHOR]
- Published
- 2008
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