Megaplasmid pKB1 of the Rubber-Degrading Bacterium Gordonia westfalica Strain Kb1
Megaplasmid pKB1 of the Rubber-Degrading Bacterium Gordonia westfalica Strain Kb1
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橡胶降解细菌 Gordonia westfalica 菌株 Kb1 的巨质粒 pKB1
DOI：
10.1007/978-3-540-85467-8_14
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发表时间：
2009
期刊：
影响因子：
--
通讯作者：
A. Steinbüchel
中科院分区：
文献类型：
--
作者：
Bröker;A. Steinbüchel
对橡胶降解菌Gordonia westfalica菌株Kb 1的环状质粒pKB 1进行序列分析，发现其含有105个开放阅读框（ORF），可分为3个功能群：（a）复制和分配，（B）催化，（c）接合转移。pKB 1的成功接合转移证明了其接合转移基因的功能。鉴定了pKB 1的复制起点，并用于构建两个适用于几种戈登氏菌和相关属的大肠杆菌戈登氏菌shuttle载体。作为pKB 1编码的cadAinE. cadA作为pKB 1特异性选择标记，通过电穿孔和接合监测pKB 1转移到分类学上相关的细菌，介导最大浓度为800 μ M的镉抗性到携带pKB 1的重组菌株。质粒pKB 1缺失突变体G. Westfalica菌株Kb1丧失了利用天然橡胶（NR）作为唯一碳源的能力，从而表明NR降解所必需的基因由pKB 1编码。对pKB 1编码的代谢功能基因的转录分析表明，在G.而在乙酸钠上生长不明显或仅有轻微的抑制作用。作为pKB 1在宿主G. Westfalicastrain Kb1由于缺乏有效的基因转移系统而失败，将pKB1转移到遗传上接近的菌株中进行工程改造和pKB1编码特征的分析。
Sequence analysis of the circular 101,016-bp plasmid pKB1 from the rubber-degrading bacteriumGordonia westfalicastrain Kb1 revealed 105 open reading frames (ORFs) which could be assigned to three functional groups (a) replication and partitioning, (b) catabolism, and (c) conjugative transfer. Successful con-jugative transfer of pKB1 demonstrated the functionality of its conjugative transfer genes. The origin of replication of pKB1 was identified and used for construction of twoEscherichia coliGordoniashuttle vectors suitable for severalGordoniaspecies and related genera. As expression of the pKB1-encodedcadAinE. colimediated resistance to cadmium,cadAwas used as a pKB1-specific selection marker to monitor transfer of pKB1 by electroporation and conjugation to taxonomically related bacteria, mediating cadmium resistance to a maximal concentration of 800 μ M to the recombinant pKB1-harboring strains. Plasmid pKB1-free mutants ofG. westfalicastrain Kb1 had lost the ability to use natural rubber (NR) as sole carbon source, thereby suggesting that genes essential for NR degradation are encoded by pKB1. Transcription analysis of pKB1-encoded genes with a putative metabolic function revealed that ORF42 (a putative cytochromecoxidase) and ORF6 (a putative epoxide hydrolase) are induced in cells ofG. westfalicastrain Kb1 during growth on NR but not or only slightly on sodium acetate. As genetic engineering of pKB1 in its hostG. westfalicastrain Kb1 failed due to the lack of an effective gene transfer system for this strain, pKB1 was transferred to genetically approachable strains for engineering and analysis of pKB1-encoded features.
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期刊：
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影响因子：
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