Performance test of cocoa’s (Theobroma cacao L.) LEAFY and AGAMOUS promoters in tobacco plant (Nicotiana tabacum L.)1)

Abstract

Performance of cocoa’s (Theobroma cacao L.) gene promoters, TcLFY and TcAG, in a tobacco plant (Nicotiana tabacum L.) used as a plant model has been tested. The two promoters were introduced into the tobacco plant cells using a modified leaf discs method employing Agrobacterium tumifaciens AGL0. The transformed tobacco plant cells were then regenerated and their morphology were observed periodically. The tobacco plants harbouring the gene promoter TcLFY, during regeneration, showed a leaf morphology similar to that of the positive control. However, the growth rate of tobacco plants inserted with promoter TcLFY was faster compared to that of the positive control. Tobacco plants harbouring gene promoter TcAG, during regeneration, showed imperfect leaf morphology. The growth rate of these tobacco plants was slower than that of the positive control and of the tobacco plants inserted harbouring gene promoter TcLFY. These results indicated that the introduced promoters affect the morphology of the tobacco plants. In order to further confirm the presence of the GFP gene and activation of TcLFY: GFP and TcAG: GFP, a molecular verification test was carried out. Variation of sucrose concentration in the media was made to examine the effect of sucrose on the activity promoters TcAG and TcLFY in tobacco plants. The sucrose concentration tested were 0%, 0,5% and 4% followed by observation on the 3rd and 7th day of cultivation. At 0% sucrose concentration the tobacco plants harbouring promoter TcLFY showed yellowish leaf, while those cultivated in the media with sucrose concentration of 0.5% and 4% grew well and normal. This indicated that sucrose affects the activity of the promoter TcLFY in tobacco plants. Tobacco plants inserted with promoter TcAG did not show significant morphological changes when grown in a medium of 0%, 0,5% or 4% sucrose concentration. This indicated that addition of sucrose does not significantly affect the induction of expression of the AG gene and the activation of the promoter TcAG.