Abstract
Jasmonate (JA) and its derivatives play an important role in plant development and defense. The JA signal receptor is an F-box protein COI1. COI1 combines with SKP1 and Cullin1 to form a SCFCOI1 complex that interacts with JAZ repressors after combining with JA leading to ubiquitination and degradation of JAZ1 by 26S proteasome. JA-related transcriptional factors are released to control the downstream gene expression. Identification of COI1 is essential to understand the JA signaling pathway. However, expression and purification of bioactive COI1 protein is extremely difficult, limiting the study of further biological function. In this study, COI1 protein was fused with specific sequence tag and transferred into tobacco. After optimizing transformation time, and extraction procedure, the fusion protein was successfully isolated by affinity purification. Pull-down assay indicated that the purified protein interacted with JAZ1 present in JA-Ile, suggesting that the fusion protein was bioactive. The methodology developed in this study provides an efficient strategy to express and purify eukaryotic protein with bioactivity.
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