| 3-PBA降解菌BA3的降解特性及基因工程菌构建 |
| 摘要点击 2859 全文点击 1808 投稿时间：2010-03-11 修订日期：2010-05-11 |
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| 中文关键词 3-苯氧基苯甲酸 生物降解 鞘脂菌 菊酯水解酶基因 基因工程菌 |
| 英文关键词 3-phenoxybenzoic acid(3-PBA) biodegradation Sphingobium sp. pyrethroid hydrolase gene(pytH) genetically engineered microorganism |
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| 中文摘要 |
| 从石油污染土壤中分离到1株能以3-苯氧基苯甲酸（3-PBA）为唯一碳源生长的细菌，命名为BA3.根据其生理生化特征和16S rDNA序列相似性分析，将该菌鉴定为鞘脂菌属(Sphingobium sp.).该菌株在60 h内对100 mg·L-1的3-苯氧基苯甲酸的降解率达到99%.降解3-苯氧基苯甲酸的最适温度为30℃，pH值为7.0，降解速率与初始接种量呈正相关.拟除虫菊酯类杀虫剂水解酶基因pytH为本实验室克隆到的一个新的菊酯水解酶基因，通过PCR从菊酯降解菌株JZ-2 总DNA 扩增了pytH，将pytH定向克隆到pBBRMCS-5载体上，构建重组质粒pPYTH，在辅助质粒pRK600的帮助下，通过三亲接合将pPYTH转移到3-PBA降解菌株Sphingobium sp.BA3 中，获得工程菌Sphingobium sp. BA3-pytH；甲氰菊酯的降解实验表明，菌株JZ-2在48 h 内对50 mg·L-1的甲氰菊酯降解率仅为60%左右，工程菌BA3-pytH相同条件下对50 mg·L-1的甲氰菊酯降解率达到95%以上，同时对甲氰菊酯的降解中间产物3-PBA也能较快降解，不会对菊酯的降解产生反馈抑制.工程菌BA3-pytH因其对甲氰菊酯和其降解中间产物3-PBA的快速降解能力，比原始菌株JZ-2在菊酯污染土壤生物修复应用中更具有优势. |
| 英文摘要 |
| A bacterium capable of utilizing 3-phenoxybenzoic acid (3-PBA) as sole carbon source was isolated from petroleum-contaminated soil. This bacterium, designated as BA3, was identified as Sphingobium sp. according to its physiological & biochemical characteristic and the similarity analysis of its 16S rDNA sequence. Strain BA3 was able to degrade 99% of 100 mg· L-1 3-phenoxybenzoic acid within 60 h. The optimal pH and temperature for the degradation were 7.0 and 30℃, respectively. The degradation efficiency was related positively to initial inoculum size. The pyrethroid hydrolase gene(pytH) gene was amplified from the genomic DNA of Sphingobium sp. JZ-2 by PCR. Recombinant plasmids pPYTH was constructed by ligating pytH gene into the broad host vector pBBRMCS-5. Under the help of plasmid RK600, pPYTH was transferred into Sphingobium sp. BA3 to construct engineering strain BA3-pytH; Fenpropathrin degradation experiments showed that strain JZ-2 was able to degrade only 60% of 50 mg·L-1 fenpropathrin in 48h while engineering strain BA3-pytH was able to degrade over 95% of 50 mg·L-1 fenpropathrin under the same conditions. Even more, BA3-pytH could rapidly degrade 3-PBA, metabolic products of pyrethroid insecticides,eliminating the inhibition of 3-PBA to pyrethroid hydrolase. Therefore, in contrast to strain JZ-2, engineering strain BA3-pytH had more advantages in bioremediation of pyrethroid insecticides contaminated environment. |
