Document Detail

Flow cytometry-based method for isolating live bacteria with meta-cleavage activity on dihydroxy compounds of biphenyl.
MedLine Citation:
PMID:  20471608     Owner:  NLM     Status:  MEDLINE    
A new method for isolating targeted live bacterial cells was established with the use of cell sorting by flow cytometry (FCM) based on the fluorescence of the intermediate metabolite of biphenyl degradation. During biphenyl degradation, a PCB degrader, Comamonas testosteroni TK102, produces a meta-cleavage intermediate metabolite, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid (HOPDA), which emits green fluorescence. HOPDA was produced from 2,3-dihydroxy biphenyl as a substrate, but it was not appropriate for labeling cells because it was released from the cells into the medium. When we used 4-n-butylbiphenyl and 4-n-heptylbiphenyl, we found that the cells produced and accumulated 2,3-dihydroxy intermediate metabolites. By the addition of synthesized 2,3-dihydroxy-4'-butylbiphenyl (2,3-DHBBP), we were able to label the cells with strong green fluorescence, suggesting the persistence of fluorescent intermediate metabolite in the cells by the introduction of the alkyl tail. 2,3-DHBBP was then used to label strain TK102 and the cells were sorted with FCM. The sorting efficiency of FCM was defined as the percentage of colony numbers per sorting events. Strain TK102 cells were successfully enriched by 4.1-fold from the mixture with environmental indigenous bacteria with a sorting efficiency of 7.3%. The method we present here serves as a basic technique for the specific and direct isolation of live bacterial cells which contain dioxygenases active on dihydroxylated aromatic compounds.
Sou Iijima; Yumi Shimomura; Yousuke Haba; Fusako Kawai; Akio Tani; Kazuhide Kimbara
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2009-12-24
Journal Detail:
Title:  Journal of bioscience and bioengineering     Volume:  109     ISSN:  1347-4421     ISO Abbreviation:  J. Biosci. Bioeng.     Publication Date:  2010 Jun 
Date Detail:
Created Date:  2010-05-17     Completed Date:  2010-07-19     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  100888800     Medline TA:  J Biosci Bioeng     Country:  Japan    
Other Details:
Languages:  eng     Pagination:  645-51     Citation Subset:  IM    
Copyright Information:
(c) 2009. Published by Elsevier B.V.
Research Institute for Bioresources, Okayama University, Kurashiki, Okayama 710-0046, Japan.
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MeSH Terms
Bacteria / enzymology,  isolation & purification*
Biphenyl Compounds / metabolism*
Comamonas testosteroni / metabolism
Dioxygenases / metabolism
Fatty Acids, Unsaturated / chemistry,  metabolism
Flow Cytometry / methods*
Polychlorinated Biphenyls / metabolism
Reg. No./Substance:
0/Biphenyl Compounds; 0/Fatty Acids, Unsaturated; 0/Polychlorinated Biphenyls; 50480-67-6/2-hydroxy-6-oxo-6-phenyl-2,4-hexadienoic acid; 92-52-4/diphenyl; EC 1.13.11.-/Dioxygenases

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