| Summary: | This study aimed to explore the influence of temperature on the growth and isoprene metabolism pathways of <i>Eucommia ulmoides</i> seedlings. Growth indices and levels of rubber and total triterpenoids in leaves were assessed under three temperature treatments: low (15 °C/10 °C), control (25 °C/20 °C), and high (35 °C/30 °C). High-throughput sequencing identified 2309 differentially expressed genes, of which 1608 were upregulated and 701 were downregulated. After 40 days, leaf length, leaf width, and plant height were significantly lower in low- and high-temperature treatments compared with the control. Specific leaf weight was higher in LT-treated leaves. Chlorophyll a and b contents were 1.372 mg.g<sup>−1</sup> and 0.594 mg.g<sup>−1</sup>, respectively, in control leaves, followed by low- and high-temperature treatments. Carotenoid content was the highest in LT treatment. The rubber content of LT and HT groups significantly decreased by 16.5% and 38%, respectively, compared with that of the control group. Total triterpene content was the lowest in control leaves at 1.02%, which was 30% and 20% less than that in low- and high-temperature treatments, respectively. This study provides insights into the efficient cultivation of <i>E. ulmoides</i> and the regulatory network of secondary metabolic pathways.
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