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粟弯孢霉叶斑病菌钙调磷酸酶基因克隆及表达分析

DOI: 10.7668/hbnxb.2013.05.020, PP. 110-115

Keywords: 新月弯孢病菌,钙调磷酸酶,qRT-PCR,环孢素A

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Abstract:

粟弯孢霉叶斑病为谷子重要的叶部病害,为了解钙调磷酸酶信号途径在粟弯孢霉叶斑病菌中的功能,对粟弯孢霉叶斑病菌钙调磷酸酶基因进行了克隆和初步分析。通过简并PCR法扩增获得了粟弯孢霉叶斑病菌钙调磷酸酶催化亚基ClCna和调节亚基ClCnb基因的同源片段,并利用RACE技术获得了ClCna和ClCnb基因序列同源片段的全长cDNA序列。粟弯孢霉叶斑病菌ClCna和ClCnb基因编码的蛋白与其他病原真菌钙调磷酸酶蛋白具有很高的同源性,ClCna基因编码的蛋白含有钙调素结合位点、钙调磷酸酶调节亚基结合位点和自身抑制调节位点,而ClCnb基因编码的蛋白含有4个钙离子结合位点。利用钙调磷酸酶特异性抑制剂环孢素A处理粟弯孢霉叶斑病菌分生孢子,发现分生孢子的萌发率明显下降。利用实时荧光定量PCR方法分析粟弯孢霉叶斑病菌中ClCna和ClCnb基因在非生物胁迫下的表达模式,在山梨醇和氯化钠胁迫下都是诱导表达,因此,推测这2个基因可能在粟弯孢霉叶斑病菌的逆境胁迫及孢子萌发中起重要的作用。

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