Source: SEAMEO BIOTROP's Research Grant | 2005
The potential endophytic fungi, Aspergillus sp. had been isolated and tested in several agriculture and forestry plants in our laboratory. A marker gene is required in studying biological aspects of the fungi for developing potential biofertilizer. The objective of this research was to integrate the GFP gene as a marker gene into the genom of endophytic fungi Apergillus sp. The GFP gene contain ToxA promoter, nos terminator and hygromycin resistance as selection marker. This study revealed that the GFP gene was successfully integrated into the endophytic fungi Aspergillus sp. using Poly Ethylene Glycol (PEG) method. Transformation efficiency of the gene integration using PEG method was 2.4% for 1µg plasmid used and germination percentage of transgenic protoplast was 48% in 36 hours incubation at room temperature. The gene integration analysis using hygromycin selection marker, fluorescence microscope under blue light filter of 515 nm wave length, and PCR using nos terminator gene indicated that the gene integration was express d constitutively in all fungal structures. The gene integration was also stable until three sub culture generation tested. The gene expression was increased by subculture treatment.
Key words: Gene transformation, GFP, endophytic fungi, biofertilizer, fungal protoplast
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