1. Enhanced Agrobacterium-mediated Transformation of Embryogenic Calli of Upland Cotton via Efficient Selection and Timely Subculture of Somatic Embryos.
- Author
-
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
- *
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 β-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. [ABSTRACT FROM AUTHOR]
- Published
- 2008
- Full Text
- View/download PDF