重寄生拟盘多毛孢cr013菌株聚酮合酶基因的多样性分析  

The Diversity Analysis of Polyketide Synthase Genes in Mycoparasite Pestalotiopsis sp.cr013 Strain

作　　者：孔磊 余进德 张登云 梅超 陈玉惠 李靖 KONG Lei;YU Jin-de;ZHANG Deng-yun;MEI Chao;CHEN Yu-hui;LI Jing(Life Science College,Southwest Forestry University,Kunming Yunnan 650224 P.R.China)

机构地区：[1]西南林业大学生命科学学院,云南昆明650224

出　　处：《西部林业科学》2021年第3期144-150,156,共8页Journal of West China Forestry Science

基　　金：云南省农业基础研究联合专项面上项目〔2017FG001(-043)〕;中央引导地方科技发展专项资金(219001);校博士科研启动金项目。

摘　　要：本研究通过基因组挖掘的方式首次从重寄生拟盘多毛孢cr013菌株中获得了聚酮合酶(polyketide synthase,PKS)基因,并利用NCBI、CDD、Clustal W、MEGA 7.0等生物信息软件对获得的PKS基因进行功能预测,挑选相似性较高、结构和功能已知的非还原型crPKS3基因和还原型crPKS12基因检测在改良Fries培养基上5个不同时间段(4、8、12、16、20 d)的基因表达情况,为重寄生拟盘多毛孢中还原型聚酮类化合物生物合成机制的阐明奠定基础。结果发现,经过全基因组数据和生物信息学分析挖掘到28个PKS(编号:crPKS 1~28)基因,包括14个HR-PKS,4个PR-PKS,8个NR-PKS和2个杂合的NRPS/PR-PKS。PKS蛋白聚类分析结果显示,推测crPKS10、crPKS25和crPKS27可能参与了美伐他汀(compactin)的生物合成;crPKS19、crPKS12、crPKS26、crPKS13、crPKS11、crPKS23、crPKS28可能参与洛伐他汀(lovastatin)生物合成;crPKS3可能参与pestheic acid的生物合成;crPKS1可能参与黑色素(melanin)的生物合成;crPKS4、crPKS8可能参与黄色分生孢子色素(yellow conidial pigment)的生物合成;crPKS20可能参与富马霉素(fumagillin)的生物合成;其余PKS蛋白序列因未聚到已知功能的分支,无法推测其催化的化合物。实时荧光定量PCR结果显示:两条PKS基因均在改良Fries培养基培养第16 d时,基因表达量最高,但crPKS12的表达量为21.72,crPKS3的表达量为12.19,crPKS12的表达量远高于crPKS3。本研究结果为重寄生拟盘多毛孢PKS基因的功能鉴定和开发利用提供理论依据,推测还原型crPKS12基因可能参与了重寄生菌cr013中聚酮的生物合成,这为该基因的克隆表达及进一步功能研究奠定了基础。In this study,polyketide genes were first obtained from the mycoparasite strain cr013 by genome mining.The function of PKS genes were predicted by NCBI,CDD,Clustal W and MEGA 7.0 softwares.The non-reducing crpks3 gene and reducing crpks12 gene with high similarity and known structure and function were selected to detect the genes expressions in 5 different time periods(4,8,12,16,20 days)in the improved Fries medium,which laid a foundation for clarifying the biosynthesis mechanism of reduceing polyketides in mycoparasite Pestalotiopsis.The results showed that 28 PKS(crPKS1-28)genes were identified with genome-whole datas and bioinformatics analysis,including 14 highly-reducing PKS(HR-PKS),4 partial-reducing PKS(PR-PKS),8 non-reducing PKS(NR-PKS)and 2 hybrid NRPS/PR-PKS.The results of PKS proteins cluster analysis showed that crPKS10,crPKS25 and crPKS27 might participate in the Compactin biosynthesis;crPKS19,crPKS12,crPKS26,crPKS11,crPKS13,crPKS23 and crPKS28 might might participate in Lovastatin biosynthesis;crPKS3 might participate in Pestheic acid biosynthesis;crPKS1 might participate in the Melanin biosynthesis;crPKS4 and crPKS8 might participate in the Yellow conidial pigment biosynthesis;crPKS20 might participate in the Fumagillin biosynthesis;and the other PKS were not clustered to the branches with known functions.The results of qPCR showed that NR-PKS crPKS3 and HR-PKS crPKS12 genes expression were the highest at the 16th day in improved Fries medium.But the expression levels of crPKS12 was 21.72 and crPKS3 was 12.19.The expression level of crPKS12 was much higher than crPKS3.This study provides a theoretical basis for the development and utilization of PKS genes from mycoparasite Pestalotiopsis.It is suggested that the HR-PKS crPKS12 gene may be involved in the polyketides biosynthesis in mycoparasite cr013 strain,which laid a foundation for the cloning and expression of the gene and further functional research.

关 键 词：重寄生 拟盘多毛孢 基因组 聚酮合酶基因 实时荧光定量PCR 

分 类 号：S718.8[农业科学—林学]