Temperature sensitivity of phenophases can reflect how and to what degree plants could tract climate change, and is related to the ability of plants to adapt to climate change. Investigating the temperature sensitivity of phenophases of different plant species could help us to identify species that are sensitive to climate change. To date, the studies about temperature sensitivity of first flowering date (FFD) mainly focused on the temperate area, and fewer studies focused on the subtropical area. We selected Guiyang City, located in subtropical China, as the study area, and analyzed the temporal changes in FFD of plants and their temperature sensitivities based on phenological observation data of 60 typical woody plants from 1980 to 2014. In addition, we evaluated the impact of the length of time series on the stability of the estimates of temperature sensitivity. The results show that: (1) Guiyang City experienced notable climate change with significantly increased annual mean temperature during the study period. The warming of spring and autumn was stronger than summer and winter. (2) FFD of 53 species (88.3%) advanced during the study period with 13 species (21.7%) significantly advancing (P<0.05). Most trends of FFD were between -4 and -2 d/decade. The overall advancing trend for FFD of 60 species was2.89 d/decade. (3) FFD was significantly and negatively correlated with mean temperature during the optimum period for most species (88.3%). Most temperature sensitivities of FFD ranged from -8 to -4 d/°C. The overall temperature sensitivity for FFD of all species was -5.75 d/°C. (4) Sample size clearly affected stability of the estimates of temperature sensitivity. The time series of 15 years could make the difference of estimates less than2 d/°C with a probability of 99%. Thus, time series should be as long as possible to be used in estimating the temperature sensitivity of flowering phenology with a stability that is sufficient for interspecific comparisons.