A 2665-year ring-width chronology was developed based on Qilian juniper from the upper treeline of the Animaqin Mountains on the eastern Tibetan Plateau. Correlation analysis results showed that the chronology was significantly negatively correlated with April–June maximum temperature at nearby meteorological stations, indicating that maximum temperature is the factor that limits tree growth in this area. Accordingly, we reconstructed the average April–June maximum temperature variations since 261 BC. Our regression model explained 37.9% of the total variance for the whole calibration period of 1960–2012. Our reconstruction revealed that the maximum temperature started to increase from approximately 1750 without a rapid warming trend, and the warmest period was from AD 890 to 947, as opposed to the recent period, whereas the period from AD 351–483 was the coldest. Significant periods in the wavelet power spectrum were approximately 2–8 years, 20–30 years, 30–60 years, and 60–130 years, as well as some long-term periods (more than 200 years). Comparisons with other temperature series from neighboring regions and the Northern Hemisphere as a whole support the validity of our reconstruction and suggest that it provides a representation of the temperature change for the Animaqin area, although asymmetric variation patterns in minimum and maximum temperatures were found.