Application of Transposon System for Selectable Marker Elimination in Transgenic Phalaenopsis
| Autor: | Yi-Ling Fan, 范藝齡 |
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| Rok vydání: | 2010 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 98 The presence of antibiotic resistant genes in genetically engineered crops together with the target gene has generated a number of environmental and consumer concerns. use of the non-antibiotic selectable marker, or elimination of marker gene from transgenic plants are suitable strategies to reduce the concerns. Plants with mutated 5-enolpyruvyl shikimate 3-phosphate synthase (EPSPS) gene are resisted to the herbicide glyphosate, which has low toxicity and non-impact on environment and animals. In order to establish a non-antibiotic selection system for Phalaenopsis transformation in this research, the modified EPSPS gene was cloned by polymerase chain reaction (PCR) from transgenic rice, ligated with CaMV 35S promoter, constructed into vector. and transformed into Phalaenopsis calli. The nature tolerance of Phalaenopsis calli against glyphosate is at the concentration of 1 mM. After transformation, the Phalaenopsis calli were selected by glyphosate and the putative transgenic calli got positive results in GUS histochemical analysis. The selection efficiency was four folds higher than G418. It suggested that glyphosate is efficient to be a selective reagent for transformed Phalaenopsis calli. Meanwhile, transposon was used as a strategy to remove selection marker. Transgenesis of Phalaenops cells and tobacco plants were performed by PCR. Transposition was confirmed after transgenic calli treated with 5 mM salicylic acid for elimination of the transposon, via PCR with suitable primers. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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