亲环素(cyclophilin)广泛存在于动植物和丝状真菌中,参与细胞信号转导,细胞凋亡,催化蛋白质折叠等过程,与病原真菌形态建成和致病过程密切相关。研究旨在获得玉米大斑病菌(Setosphaeria turcica)亲环素基因,通过表达规律分析明确其与病菌生长发育的关系。利用简并引物PCR结合RACE技术获得S. turcica中亲环素基因的全长,并通过Real-time PCR技术检测该基因在病菌侵染结构发育过程中的表达模式。结果表明,玉米大斑病菌亲环素基因开放阅读框全长1125 bp,3"UTR 154 bp,5"UTR 93 bp,编码374个氨基酸,将此基因命名为CyPs1,并将其cDNA序列提交GenBank,获得登录号EU679371.1,Protein ID为ACD62431.1。系统发育树分析显示CyPs1与玉米小斑病菌、蓝莓枯枝病菌等物种的亲环素同源性可达到90%以上。该基因在病菌分生孢子萌发、附着胞形成及侵染阶段均有表达,至附着胞形成和侵染菌丝形成阶段,转录水平分别升至分生孢子时期的2倍和3倍。该研究为进一步了解玉米大斑病菌Ca2+信号途径的功能及作用机制奠定了基础。
英文摘要:
Cyclophilin, as a highly conserved protein family, is widely found in plants, aminals and filamentous fungi, and is involved in cell signaling, apoptosis, protein folding process, and so on. It also closely relates to morphogenesis and pathogenicity of pathogenic fungi. The aim of this study is to obtain the cyclophilin gene and clarify the mechanism of cyclophilin regulating the morphogenesis and pathogenicity of Setosphaeria turcica. The full length of the cyclophilin gene was obtained through degenerate primer PCR combined with RACE technique and its expression patterns were detected at different developmental stages based on quantitative real-time PCR (qRT-PCR) analysis. The results were as follows: the full-length open reading frame (ORF) of the cyclophilin gene was 1125 bp, with a 3"UTR of 154 bp and a 5"UTR of 93 bp, encoding a protein of 374 amino acids. The gene was named CyPs1 and its cDNA sequence was submitted to GenBank with the accession number EU679371.1 and Protein ID ACD62431.1. Phylogenetic tree analysis showed that it was associated with the homologues in Bipolaria maydis and Neofusicoccum parvum, and the homology was more than 90%. The expression pattern showed that CyPs1 was persistantly expressed during the developmental process of invasive structures. Its transcription level in the stage of appressorium formation and invasive mycelial morphogenesis increased to 2 and 3 times of that of conidia, respectively. This study laid the foundation for further understanding of the function and mechanism of Ca2+ signaling in S. turcica.
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