关键词: 金鱼草, Expansin基因家族, 表达模式, 核盘菌抗性, 抗性初步筛选

Abstract:

Objective Investigate the role of expansin (EXPs) of snapdragon (Antirrhinum majus) in response to Sclerotinia sclerotiorum infection, identify the EXPs gene family of snapdragon, and mine the key candidate genes against S. sclerotiorum. It provides a basis for further investigation of the biological function of the EXPs gene family in snapdragon. Method Based on the whole snapdragon genome data, bioinformatics methods were used to analyze the physicochemical properties, secondary and tertiary structures, gene structure, protein domains, phylogenetic evolution, collinearity relationships and cis-acting elements of the EXPs family proteins in snapdragon. The transcriptome sequencing (RNA-seq) and quantitative real-time PCR (RT-qPCR) were applied to examine the expression patterns of AmEXPs during S. sclerotiorum infection while using resistant and susceptible A. majus materials. Candidate genes were identified. These genes were further investigated by constructing overexpression vectors and transiently transformed into Nicotiana benthamiana for preliminary resistance screening. Result Thirty-two AmEXPs were identified in the snapdragon, all of them have conserved structural domains DPBB-1 and Pollen allerg1, and they are divided into four subfamilies (EXPA, EXPB, EXLA, and EXLB). Thy encoded 211-297 amino acids, with molecular weights of 22 018.69-31 787.97 Da, and isoelectric points ranging from 5.01-9.83. There are highly similar structures and conserved motifs among the genes of various subfamilies. Each gene's promoter contains cis-acting elements related to hormones or stress. Ten candidate genes were identified according to their expression patterns. These genes were further investigated by constructing overexpression vectors and transiently transformed into N. benthamiana for preliminary resistance screening. It was found that AmEXP13 and AmEXLB1 genes significantly enhanced the resistance of N.benthamiana to S. sclerotiorum compared to the control, while AmEXPA17, AmEXPA21, AmEXPA9, and AmEXPA16 genes made N. benthamiana more susceptible, and AmEXLB2, AmEXPB1, AmEXPA2, and AmEXPA12 genes showed no significant differences. Conclusion Thirty-two EXPs gene family members were identified from the snapdragon genome, among which six EXPs genes respond to resistance to S. sclerotiorum. AmEXPA13 and AmEXLB1 are the key candidate genes for positive regulation of resistance, while AmEXPA17, AmEXPA21, AmEXPA9, and AmEXPA16 are the key candidate genes for negative regulation of resistance.

Key words: Antirrhinum majus, Expansin gene family, expression patterns, resistance to Sclerotinia sclerotiorum, preliminary resistance screening