Transformation of Maize Cells and Regeneration of Fertile Transgenic Plants.
W. J. Gordon-Kamm,T. M. Spencer,Mary L. Mangano,Thomas R. Adams,Richard J. Daines,William G. Start,James V. O'brien,Sheryl A. Chambers,W. R. Adams,Nancy Willetts,Thomas B. Rice,Catherine J. Mackey,R. W. Krueger,Albert P. Kausch,Peggy G. Lemaux +14 more
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TLDR
A reproducible system for the generation of fertile, transgenic maize plants has been developed and activity of the enzyme phosphinothricin acetyltransferase (PAT) encoded by bar were confirmed in all bialaphos-resistant callus lines.Abstract:
A reproducible system for the generation of fertile, transgenic maize plants has been developed. Cells from embryogenic maize suspension cultures were transformed with the bacterial gene bar using microprojectile bombardment. Transformed calli were selected from the suspension cultures using the herbicide bialaphos. Integration of bar and activity of the enzyme phosphinothricin acetyltransferase (PAT) encoded by bar were confirmed in all bialaphos-resistant callus lines. Fertile transformed maize plants (R0) were regenerated, and of 53 progeny (R1) tested, 29 had PAT activity. All PAT-positive progeny analyzed contained bar. Localized application of herbicide to leaves of bar-transformed R0 and R1 plants resulted in no necrosis, confirming functional activity of PAT in the transgenic plants. Cotransformation experiments were performed using a mixture of two plasmids, one encoding PAT and one containing the nonselected gene encoding [beta]-glucuronidase. R0 plants regenerated from co-transformed callus expressed both genes. These results describe and confirm the development of a system for introduction of DNA into maize.read more
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References
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A revised medium for rapid growth and bio assays with tobacco tissue cultures
Toshio Murashige,Folke Skoog +1 more
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A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity
TL;DR: In this article, a technique for conveniently radiolabeling DNA restriction endonuclease fragments to high specific activity is described, where DNA fragments are purified from agarose gels directly by ethanol precipitation and are then denatured and labeled with the large fragment of DNA polymerase I, using random oligonucleotides as primers.
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