Construction of a dual fluorescence whole-cell biosensor to detect N-acyl homoserine lactones

Detection of N-acyl homoserine lactones （AHLs） is useful for understanding quorum sensing （QS） behaviors, including biofilm formation, virulence and metabolism. For detecting AHLs and indicating the host cells in situ, we constructed the plasmid pUCGMA2T1-4 to make a dual fluorescent whole- cell bio...

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Bibliographic Details
Published inJournal of environmental sciences (China) Vol. 26; no. 2; pp. 415 - 422
Main Authors Deng, Xuemei, Zhuang, Guoqiang, Ma, Anzhou, Yu, Qing, Zhuang, Xuliang
Format Journal Article
LanguageEnglish
Published Netherlands Elsevier B.V 01.02.2014
Subjects
4-Butyrolactone - analogs & derivatives
4-Butyrolactone - analysis
Biosensing Techniques
dual fluorescence
Escherichia coli
Fluorescence
gfp
indicator
mcherry
N-酰基高丝氨酸内酯
Plasmids
Pseudomonas syringae
Quorum Sensing
whole-cell biosensor
信号分子
全细胞
双荧光
宿主细胞
检测
生物传感器
荧光响应
gfp
indicator
quorum sensing
whole-cell biosensor
mcherry
dual fluorescence
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Summary:Detection of N-acyl homoserine lactones （AHLs） is useful for understanding quorum sensing （QS） behaviors, including biofilm formation, virulence and metabolism. For detecting AHLs and indicating the host cells in situ, we constructed the plasmid pUCGMA2T1-4 to make a dual fluorescent whole- cell biosensor based on the AhlI/R AHL system of Pseudomonas syringae pv. syringae B728a. The plasmid contains three components： constitutively expressed enptll：：gfP for indicating host cells, Pahll：：mcherry that produces red fluorescence in response to AHL, and the ahIR gene that encodes an AHL regulatory protein. Meanwhile, two copies of T1-4 （four tandem copies of a transcriptional terminator） were added into the plasmid to reduce background. The results showed that when the plasmid was placed into Escherichia coli, the dual fluorescence whole-cell biosensor was able to respond with red fluorescence within 6 hr to 5 × 10^-8-1 × 10^-5 mol/L of 3OC6-HSL. Bright green fluorescence indicated the host cells. Furthermore, when the plasmid was transferred to wild- type Pseudomonas PhTA125 （an AHL-producing bacterium）, it also showed both green and red fluorescence. This result demonstrates that this plasmid can be used to construct whole-cell indicators that can indicate the AHL response and spatial behaviors of microbes in a mi tal niche
Bibliography:whole-cell biosensor quorum sensing dual fluorescence gfp mcherry indicator
Detection of N-acyl homoserine lactones （AHLs） is useful for understanding quorum sensing （QS） behaviors, including biofilm formation, virulence and metabolism. For detecting AHLs and indicating the host cells in situ, we constructed the plasmid pUCGMA2T1-4 to make a dual fluorescent whole- cell biosensor based on the AhlI/R AHL system of Pseudomonas syringae pv. syringae B728a. The plasmid contains three components： constitutively expressed enptll：：gfP for indicating host cells, Pahll：：mcherry that produces red fluorescence in response to AHL, and the ahIR gene that encodes an AHL regulatory protein. Meanwhile, two copies of T1-4 （four tandem copies of a transcriptional terminator） were added into the plasmid to reduce background. The results showed that when the plasmid was placed into Escherichia coli, the dual fluorescence whole-cell biosensor was able to respond with red fluorescence within 6 hr to 5 × 10^-8-1 × 10^-5 mol/L of 3OC6-HSL. Bright green fluorescence indicated the host cells. Furthermore, when the plasmid was transferred to wild- type Pseudomonas PhTA125 （an AHL-producing bacterium）, it also showed both green and red fluorescence. This result demonstrates that this plasmid can be used to construct whole-cell indicators that can indicate the AHL response and spatial behaviors of microbes in a mi tal niche
11-2629/X
ObjectType-Article-1
SourceType-Scholarly Journals-1
ObjectType-Feature-2
content type line 23
ISSN:1001-0742
1878-7320
DOI:10.1016/S1001-0742(13)60407-6