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Bioremediation of xenobiotic compounds including dioxins

包括二恶英在内的外源化合物的生物修复

基本信息

批准号：
11794005
负责人：
OMORI Toshio
金额：
$ 5.76万
依托单位：
THE UNIVERSITY OF TOKYO
依托单位国家：
日本
项目类别：
Grant-in-Aid for University and Society Collaboration
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11794005/
关键词：
Dioxin Bioremediation Bioaugmentation Gene transfer Phytoremediation Monitoring Transgenic Plants Phytoremedhiation

项目摘要

[1] Two kinds of bacteria, a dinenzofuran (DF)-degrader Terrabacter sp. strain DBF63 and a carbazole (CAR)-degrader Pseudomonas resinovorans strain CA10, were investigated for their ability to transform several chlorinated DFs and dibenzo-p-dioxins (DDs). It was shown that CAR 1,9a-dioxygenase from strain CA10 catalyzed angular dioxygenation of all mono- to tri-chioro-DF/DDs (CDF/CDDs) investigated in this study, but DF 4,4a-dioxygenase from strain DBF63 did not transform 2,7-diCDD. We also tried a preliminary bioremediation of the actual dioxin-contaminated soil using the soil-slurry system. The decrease of total CDD and CDF congeners by single inoculation with CA10 or DBF63 cells were 8.3 and 10%, respectively, after 7-day incubation, thus it was shown that strains CA10 and DBF63 had a potential to degrade tetra- to hepta-chlorinated congeners including the most toxic compound, 2,3,7,8-tetraCDD.[2] The fluorogenic probe assay, competitive PCR and co-extraction with internal standard cells were combined to develop a rapid, sensitive, and accurate quantification method for the copy number of a target carAa gene and the cell number of strain CA10. In addition, by using Tn5 transposon, strain CA10 was chromosomally marked with a tandem gfp gene. The real-time competitive PCR and direct counting using the GFP marker were employed to monitor the total number of carAa gene and survival of strain CA10 cells in the soil.[3] Following plate matings between CAR-degrader strain K23 and phenol-degrader strain DS1 in the presence of CAR, transconjugants able to utilize CAR were obtained. The transconjugants had large plasmid pCAR2.[4] Transgenic plants expressing DbfB fused to endoplasmic reticulum signal peptide continuously secrete recombinant proteins from their roots into a simple hydroponic medium, and the secreted DbfB proteins retained biological activity.

[1]研究了2种细菌对氯代二苯并呋喃（DF）和二苯并二恶英（DDs）的降解能力，分别为Terrabacter sp.菌株DBF 63和Pseudomonasresinovorans菌株CA 10。结果表明，来自菌株CA 10的CAR 1，9 α-双加氧酶催化本研究中研究的所有单至三氯-DF/DD（CDF/CDD）的角向双氧化，但来自菌株DBF 63的DF 4，4 α-双加氧酶不转化2，7-二CDD。我们还尝试了初步的生物修复的实际二恶英污染的土壤使用土壤泥浆系统。单次接种CA 10或DBF 63细胞培养7天后，总CDD和CDF同系物的减少分别为8.3%和10%，因此表明菌株CA 10和DBF 63具有降解四氯至七氯同系物的潜力，包括毒性最大的化合物2，3，7，8-tetraCDD。[2]将荧光探针法、竞争PCR法和内标细胞共提取法相结合，建立了一种快速、灵敏、准确的检测CA 10菌株carAa基因拷贝数和细胞数的方法。另外，利用Tn 5转座子对CA 10菌株进行了串联gfp基因的染色体标记。采用实时竞争PCR和GFP标记直接计数法检测土壤中carAa基因的表达量和菌株CA 10的存活率。[3]在CAR降解菌株K23和苯酚降解菌株DS 1之间在CAR存在下进行平板交配后，获得能够利用CAR的转移缀合物。转化接合物具有大质粒pCAR 2。[4]表达与内质网信号肽融合的DbfB的转基因植物从其根部持续分泌重组蛋白到简单的水培培养基中，并且分泌的DbfB蛋白保留生物活性。