Crop yield and quality are often limited by the amount of phosphate fertilizer added to infertile soils, a key limiting factor for sustainable development in modern agriculture. The polyphosphate kinase (ppk) gene -expressing transgenic rice with a single-copy line (ETRS) is constructed to improve phosphate fertilizer utilization efficiency for phosphorus resource conservation. To investigate the potential mechanisms of the increased biomass in ETRS in low phosphate culture, ETRS was cultivated in a low inorganic phosphate (Pi) culture medium (15 μmol/L Pi, LP) and a normal Pi culture medium (300 μmol/L Pi, CP), respectively. After 89 d of cultivation in different concentrations of phosphate culture media, the total phosphorus, polyphosphate (polyP), biomass, photosynthetic rate, nonstructural carbohydrate (NSC) contents, related enzyme activities, and related gene expression levels were analyzed. The results showed that ETRS had a high polyP amount to promote the photosynthetic rate in LP, and its biomass was almost the same as the wild type (WT) in CP. The NSC content of ETRS in LP was higher than that of WT in LP, but slightly lower than that of WT in CP. PolyP notably promoted the sucrose phosphate synthase activities of ETRS and significantly down-regulated the expression levels of sucrose transporter genes (OsSUT3 and OsSUT4), resulting in inhibiting the transport of sucrose from shoot to root in ETRS. It was concluded that polyP can stimulate the synthesis of NSCs in LP, which improved the growth of ETRS and triggered the biological activities of ETRS to save phosphate fertilizer. Our study provides a new way to improve the utilization rate of phosphate fertilizer in rice production.