Functional Analysis Of GDP-D-mannose Pyrophosphorylase(GMPase) Gene In Tomato

L-ascorbate (Vitamin C; AsA), one of the best known of the plant metabolites, is a major antioxidant that serves many functions in plants and has many proposed roles in plant growth and metabolism. Humans can not synthesize ascorbic acid because of the lack of L-gulono-1,4-lactone oxidoreductase, which is required for the final step in AsA synthesis. Therefore, AsA contents is one of the important characterised components to the nutritive value of vegetables and fruits. GDP-D-mannose pyrophosphorylase is an enzyme in initial committed step of AsA biosynthesis (L-galactose pathway). Tomato (Solanum esculentum L.)is ranking the top in the production area and total yield among all vegetable crops in the world. Therefore, in the present study, we transformed tomato with A. tumefaciens (LBA4404) carrying PBI121- GMPase to get the transgenic plant with GMPase.Further more, we studied the effect of overexpression, with a view to improving the AsA content of tomato. The results showed as fllows:1 Tomato was transformed with A. tumefaciens (LBA4404). Three transgene plants were identified by PCR amplification in the gaining six regenerated plants.2 Western bloting analysis for the three transgenic lines with the antiserum against GMPase revealed the presence of a 43 KD protein corresponding to GMPase in GMPase overexpressing plants and WT plant. Signals were obvious in two transgenic lines, it's showed that GMPase had been transfered to tomato.3 Compared with the control, the transgenic lines had significant changes in the GMPase activity which were increased about 127 and 143% respectively; the contents of AsA were also increased about 179.6 -288.9% in functional leaf and 42.3-55.6% in riping fruit, respectively, but minor change in mature green fruit.4 The enzymes (APX, AAO, DHAR, MDHAR, GR) related with AsA metabolism had different significant changes in the T1 transgenic lines, APX, AAO, DHAR, MDHAR and GR activities were increased about 77.1% and 73.9%, 24.3% and 25.6%, 38.4% and 25.3%, 46.8% and 57.1%, 57.6% and 36.4%, respectively. GSH and GSSG contents also increased, which were increased about 34.9% and 31.7%, 6.7% and 12.0%.5 The fruits of the two transgenic plants had lower acid content and had higher sugar content and sugar/acid ratio than that of control.