拟康宁木霉T-51几丁质酶活性及内切几丁质酶基因克隆与分析

摘要

Trichoderma koningiopsis T-51 is a potential biological control agent against Botrytis cinerea.In this study,chitinase activity of T-51 was significantly induced by B.cinerea in both liquid PDB medium and the interacting area with B.cinerea colony on PDA medium.An endo-chitinase gene Tkchit42 was cloned from T.koningiopsis T-51.The full length of Tkchit42 was 1 817 bp,encoding an endo-chitinase protein containing 424 amino acid residues.The predicted protein molecular weight was 46.378 kDa.The Tkchit42 amino acid sequence BLAST showed 99％ similarity with that of a 42 kDa endo-chitinase of T.koningii.For our knowledge,this is the first report of chitinase gene cloned from T.koningiopsis.%木霉是一类重要的生防真菌,木霉产生的凡丁质酶在其生物防治的重寄生过程中起着重要作用.拟康宁木霉Trichoderma koningiopsis T-51是一株对番茄灰霉病有生防潜力的木霉菌株,本文测定了T-51菌株产生几丁质酶的活性,结果表明T-51在PDB中液体培养以及与灰霉病菌在PDA上对峙培养时产生的几丁质酶活性显著受到灰霉病菌的诱导.采用RACE技术首次克隆了拟康宁木霉T-51中一个几丁质酶基因Tkchit42,全长为1 817 bp,包含4个外显子和3个内含子.预测该基因有一个1 275 bp的开放阅读框,编码424个氨基酸,预测的蛋白总分子量为46.378 kDa,预测的等电点(pI)为5.16,与T.koningii中42 kDa内切几丁质酶的氨基酸序列相似性达到99％.