Abstract: To further study the function of geranylgeranyl pyrophosphate synthase (GGPPS) gene from Nicotiana tabacum, NtGGPPS1 was successfully cloned from tobacco cultivar K326 using gene-specific primers which were designed according to the coding DNA sequence of GGPPS1(GenBank:GQ911583.1) from Nicotiana tabacum. Fluorescent expression vector of NtGGPPS1(PFF19-NtGGPPS1-GFP) was constructed for subcellular localization in tobacco. After treated with different phytohormones, the expression level of NtGGPPS1 in K326 was studied by real-time PCR. RNAi vector pHellsgate2-NtGGPPS1 was introduced into K326 via Agrobacterium-mediated transformation and the contents of plastid pigments in NtGGPPS1 down-regulated tobacco plants were detected. The results showed that green fluorescence protein NtGGPPS1-GFP fusion was distributed in the plastids of transgenic BY-2 cells, suggesting that NtGGPPS1 located in plastids. The expression level of NtGGPPS1 in K326 was significantly increased after MeJA treatment, while it was contrary to IAA treatment. Comparing with the control, the contents of plastid pigments significantly decreased in NtGGPPS1 down-regulated plants, which indicated that NtGGPPS1 was involved in the biosynthesis of β-carotene in Nicotiana tabacum.