Evaluation of Green Fluorescence Protein (GFP) as a Selectable Marker for Oil Palm Transformation via Transient Expression

Abstract. Green fluorescent protein (GFP) can be easily visualized under UV/blue light without any additional substrate or co-factor. Its assay is also non-destructive. Therefore, it has been widely used to monitor transgene expression and protein localization in variety of cells and organisms. To evaluate the effectiveness and practicality of GFP as a reporter and selectable marker for oil palm transformation, transient expression of GFP genes in oil palm embryogenic calli and immature embryos were analyzed after bombardment with different types (version) of GFP genes and driven by different constitutive promoters. Some of the GFP genes used were targeted to specific organelles within the cell, namely mitochondria, plastid and endoplasmic reticulum (ER). For transformation using non-targeted GFP genes, plasmid p35S.TYG.GFP showed the highest number of green fluorescent spots, followed by pHBT-sGFPS65T and pGEM.Ubi-sGFPS65T. The results demonstrated that the sGFPS65T gene version is the most active in oil palm. When organelle targeted GFP genes were transformed, localization of the genes into specific organelle was clearly demonstrated using laser scanning confocal microscopy. Using the organelle targeted GFP genes; the sGFPS65T gene version was again demonstrated to be the most active in oil palm cells. Long term transient expression of the GFP genes was also evaluated and is discussed