GENETIC TRANSFORMATION OF BANANA CV. AMBON KUNING’S EMBRYO CELL SUSPENSION CULTURE MEDIATED BY Agrobacterium tumefaciens

• Main Author: Magfirah Hala, Dian
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/34243
• Institution: Institut Teknologi Bandung

Quality improvement of banana is required in order to produce banana plants that resist to biotic and abiotic stress. However, quality improvement of banana by using conventional approach is generally difficult, due to multiploids and parthenocarpy characteristics. Thus, genetic transformation approach using Agrobacterium tumefaciens should be tested as an alternative solution. In this experiment, we used embryogenic cell suspension culture of banana cv. Ambon Kuning to avoid the formation of chimera at the transformed plant tissue. The objective of this research was to establish the transformation method of embryogenic cell suspension culture of banana mediated by A. tumefaciens with green fluorescent protein (GFP) as the reporter gene and nptII as the molecular marker. Embryogenic single cell suspension culture of banana was obtained from immature male flower, and then transformed by A. tumefaciens strain AGL-1 harboring pART-TEST7 plasmid (with optical density OD600 = 1) using assisted co-centrifugation method. The suspension culture was then cocultivated in semisolid media. The results of transformation were evaluated by observation of GFP gene expression in the cell suspension culture of transformed banana plant embryo using fluorescence microscope and by PCR test. The results showed that the transformation was successfully achieved confirmed by a more intense green color in the microscope observation compared to the untransformed control explant. Subsequent, PCR test also showed a band of 659 base pairs correspond to nptII gene which was integrated in the genome of transformed embryogenic cell suspension culture. Based on these results, it can be concluded that transient genetic transformation in embryogenic cell suspension culture of cv. Ambon Kuning was successfully conducted. The developed protocol can be further applied as genetic transformation protocol in local banana with certain specific genes, like pest and disease resistant genes, in order to improve the quality of banana plants in Indonesia.