Abstract: Five important genes coding for nitrate reductase (NR), nitrite reductase (NiR), glutamine synthetase (GS), glutamate dehydrogenase (GDH) and asparagine synthetase (AS), were cloned from the related EST sequences deposited in VitisEST database and amplified using RT-PCR. Sub-cellular localization was employed to characterize their functional position, while semi-quantitative RT-PCR and fluorescent quantitative PCR were both used to analyze their expression in leaves after treatment with urea at different concentrations. Results show that the five genes had significantly higher expression in younger leaves compared to old leaves. The differences in expression of these five genes at different times were inconsistent, but generally expression was increased by application of foliar urea, with application of 0.3% and 0.5% urea causing significant increases in gene expression compared to control plants. The expression of the five genes related to foliar fertilizer application was corroborated by the significant increases in related fruit indicators. Expression of GS in grape leaves had the most significant increase at 6 h after spraying of urea, while NR maintained a relatively high level of expression even at 48h after urea application compared to the other genes. NR and GS expression in various stages was higher than NiR and AS respectively, with the trends in expression of NiR and AS being consistent with those of NR and GS respectively. This study provides a theoretical basis for further studies on the mechanisms of nitrogen metabolic pathways and scientifically informed foliar fertilizer application.