Agrobacterium-mediated Transformation of gus and mgfp Reporter Genes

Abstract

Certain factors affecting transformation efficiency of teak was investigated. Two strains of Agrobacterium
tumefaciens, EHA 105 and AGL1 harboring plasmid pCAMBIA 1304 were used. It was found that
A. tumefaciens strain EHA105 with OD600 =1.0, inoculation period of 3 hours and co-cultivation period of 3
days resulted the highest efficiency of transformation and transient expression of gus reporter gene.
The transformation of node explants yielded better result than using leaf base explants. These conditions
were used to transform hygromycin phosphotransferase (hpt) gene and membrane bound green
fluorescence protein (mgfp) reporter gene into in vitro shoot apices, node with developing shoot, and
callus tissue. Two weeks after selection on 50 mg/L hygromycin containing media, the transient expression
of mgfp gene was visualized. The highest percentage of mgfp-positived tissue was found in the shoot
apices (90%) followed by node with developing shoot (73%) and callus (33%) respectively. The expression
of mgfp was found in shoot tip and young leaf. However, only 2 pieces of node explants showed stable
expression of mgfp gene.