张鹏,刘盼,周兰,裴婷,甘喆,李浩东,宋发军.枝状枝孢霉MD2的候选基因A00981的克隆及转基因菌株的构建[J].中南民族大学学报自然科学版,2019,38(1):45-49
枝状枝孢霉MD2的候选基因A00981的克隆及转基因菌株的构建
Cloning of the Candidate Gene A00981 of Cladosporium cladosporioides MD2 and Construction of A00981-transgenic Strains
  
DOI:10.12130/znmdzk.20190108
中文关键词: 枝状枝孢霉MD2  紫杉醇  基因克隆  转基因菌株
英文关键词: Cladosporium cladosporioides MD2  taxol  gene clone  transgenic strain
基金项目:国家自然科学基金资助项目(31370118, 31770134)
作者单位
张鹏,刘盼,周兰,裴婷,甘喆,李浩东,宋发军* 中南民族大学 生命科学学院 生物技术国家民委重点实验室/武陵山区特色资源植物种质保护与利用湖北省重点实验室武汉 430074 
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中文摘要:
      目的:为揭示真菌紫杉醇合成分子机制.方法:克隆了产紫杉醇内生真菌枝状枝孢霉MD2的候选基因A00981,该基因DNA和cDNA序列分别为969 bp和894 bp;其编码蛋白含有298个氨基酸、无跨膜结构和信号肽,与酰基辅酶A脱氢酶(ACAD)具有73%的一致性.进一步构建该基因的超表达载体pTFC-A00981,并转化枝状枝孢霉MD2,共获得9株转基因菌株. 结果:Southern blot分析证明其中5株转基因菌株的外源基因是以单拷贝方式插入.结论:该结果为深入分析超表达候选基因A00981对枝状枝孢霉MD2紫杉烷类合成的影响奠定了基础.
英文摘要:
      Objective: To uncover the molecular mechanism of the biosynthesis of taxol in fungi. Methods: The candidate gene A00981 of taxol-producing endophytic fungus Cladosporium cladosporioides MD2 was cloned, and the DNA and cDNA sequences of the gene were 969 bp and 894 bp, respectively. The encoded protein contained 298 amino acids, no transmembrane structure and signal peptide, and had 73% consistency with acyl-CoA dehydrogenase (ACAD). The super-expression vector pTFC-A00981 was further constructed and transformed into C. cladosporioides MD2, and nine transgenic strains were obtained. Results: Southern blot analysis showed that one copy of the exogenous gene was inserted into the genome among the 5 transgenic strains. Conclusion: The results of this study lay a foundation for further analysis of the effect of overexpression candidate gene A00981 in C. cladosporioides MD2 for taxanes synthesis.
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