Photoperiod and temperature are two pivotal regulatory factors of plant flowering. The floral transition of plants depends on accurate measurement of changes in photoperiod and temperature. The flowering time of rice (Oryza sativa) as a facultative short-day (SD) plant is delayed under long-day (LD) and/or low temperature conditions. To elucidate the regulatory functions of photoperiod and temperature on flowering time in rice, we systematically analyzed the expression and regulation of several key genes (Hd3a, RFT1, Ehd1, Ghd7, RID1/Ehd2/OsId1, Se5) involved in the photoperiodic flowering regulatory pathway under different temperature and photoperiod treatments using a photoperiod-insensitive mutant and wild type plants. Our results indicate that the Ehd1-Hd3a/RFT1 pathway is common to and conserved in both the photoperiodic and temperature flowering regulatory pathways. Expression of Ehd1, Hd3a and RFT1 is dramatically reduced at low temperature (23°C), suggesting that suppression of Ehd1, Hd3a and RFT1 transcription is an essential cause of delayed flowering under low temperature condition. Under LD condition, Ghd7 mRNA levels are promoted at low temperature (23°C) compared with normal temperature condition (28°C), suggesting low temperature and LD treatment have a synergistic role in the expression of Ghd7. Therefore, upregulation of Ghd7 might be a crucial cause of delayed flowering under low temperature condition. We also analyzed Hd1 regulatory relationships in the photoperiodic flowering pathway, and found that Hd1 can negatively regulate Ehd1 transcription under LD condition. In addition, Hd1 can also positively regulate Ghd7 transcription under LD condition, suggesting that the heading-date of rice under LD condition is also regulated by the Hd1-Ghd7-Ehd1-RFT1 pathway.