Construction of Yeast Two-Hybrid Library of Arabidopsis thalianaand Screening of ADPG1 Interacting Proteins

HAN Xue^1,2, WANG Shufang³, LIN Jinxing^1,2*

(¹College of Biological Sciences and Technology, Beijing Forestry University, Beijing 100083, China; ²National Engineering Laboratory for Tree Breeding, Beijing Forestry University, Beijing 100083 China; ³Institute of Botany, the Chinese Academy of Sciences, Beijing 100093, China)

Abstract:

Cellulase, hemicellulase and pectinase play critical roles in the fruit development and ripening. ADPG1 (ARABIDOPSIS DEHISCENCE ZONE POLYGALACTURONASE 1) is an essential pectinase for silique dehiscence. In addition, the ectopic expression of ADPG1 in xylem can induce the expression of PR (pathogenesisrelated) gene in Arabidopsis ccr1 mutant. However, the mechanism of how ADPG1 promotes the release of elicitors in ccr1 mutant and induces the expression of defense-related gene is still unclear. To explore the role of ADPG1 involving in the release of elicitors and the induction of defense gene PR1 in ccr1 mutants, the interacting proteins with ADPG1 were screened. In this paper, the authors extracted the total RNA from wild-type and ccr1 Arabidopsis. Then, the extracted RNA was used to isolate mRNA and synthesize double-stranded cDNA. The primary and secondary libraries of yeast two-hybrid were constructed successively. The bait vector pGBKT7-AtADPG1 was constructed and tested for its self-activation. The pGBKT7- AtADPG1 was co-transformed into yeast competent cells with the secondary library, and a candidate protein named AtGRP5 that interacting with ADPG1 was identified. This study provided a basis for further study on the involvement of ADPG1 in plant defense response.

CSTR: 32200.14.cjcb.2022.06.0009