Homologous expression of Phanerochaete chrysosporium manganese peroxidase, using bialaphos resistance as a dominant selectable marker
Homologous expression of Phanerochaete chrysosporium manganese peroxidase, using bialaphos resistance as a dominant selectable marker
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DOI：
10.1007/s00294-003-0418-z
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发表时间：
2003-07
期刊：
影响因子：
2.5
通讯作者：
B. Ma;M. Mayfield;M. Gold
中科院分区：
文献类型：
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作者：
B. Ma;M. Mayfield;M. Gold
锰过氧化物酶(MnP)是白腐菌Phanerochaete chrysosporium木质素降解系统的主要胞外成分。重组MNP同工酶rMnP1 INP的同源表达利用吸水链霉菌抗Bialaphos基因bar作为选择标记，建立了一种新的黄孢子菌转化体系。该系统的转化频率约为每微克DNA 100个比拉磷抗性转化子。转化的菌株都含有质粒DNA，异位整合到真菌基因组中。利用该转化体系，将THp基因启动子区域。利用黄孢子菌翻译延伸因子基因在原代代谢培养物OFP中驱动编码MnP1的mnp1的表达。黄孢子菌，内源MNP不表达。在转化子的搅拌培养中，每升胞外培养基产活性的重组MnP1约为2-3 mg。
Manganese peroxidase (MnP) is a major extracellular component of the lignin-degrading system of the white-rot fungus,Phanerochaete chrysosporium. Homologous expression of recombinant MnP isozyme 1 (rMnP1) inP. chrysosporiumwas achieved using a novel transformation system for this fungus, which utilizes theStreptomyces hygroscopicusbialaphos-resistant gene,bar, as the selectable marker. The transformation frequency for this system is approximately 100 bialaphos-resistant transformants per microgram of plasmid DNA. Transformed strains all contain plasmid DNA, ectopically integrated into the fungal genome. Using this transformation system, the promoter region of theP. chrysosporiumtranslation elongation factor gene was used to drive expression ofmnp1, encoding MnP1, in primary metabolic cultures ofP. chrysosporium, where endogenous MnP was not expressed. Approximately 2–3 mg of active recombinant MnP1 per liter of extracellular medium was produced in agitated cultures of transformants.