Method for measuring content of total acyl homoserine lactone (AHL)

A culture medium, the technology directly below, applied in the field of determination of total acyl homoserine lactone content, can solve the problems of time-consuming, high cost and the like

Active Publication Date: 2016-12-07
RES CENT FOR ECO ENVIRONMENTAL SCI THE CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Common chemical assay methods are time consuming and costly

Method used

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  • Method for measuring content of total acyl homoserine lactone (AHL)
  • Method for measuring content of total acyl homoserine lactone (AHL)
  • Method for measuring content of total acyl homoserine lactone (AHL)

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Experimental program
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Effect test

Embodiment 1

[0049] Embodiment 1, a kind of method for measuring acyl homoserine lactone (AHL)

[0050] In this example, Agrobacterium tumefaciens A136 is used as the AHL reporter strain, and Agrobacterium tumefaciens A136 contains the β-galactosidase lacZ reporter gene. When Agrobacterium tumefaciens A. tumefaciens A136 is exposed to exogenous AHL, its lacZ reporter gene is expressed, and its expression product can hydrolyze the substrate 5-bromo-4-chloro-3-indole-β-D-galactoside (X -gal), resulting in blue spots, different AHL concentrations have different color distances, and the color distance is positively correlated with the AHL concentration. Use Agrobacterium tumefaciens A. tumefaciens A136 to measure the color development distance of a series of standard AHL preparations with different concentrations, and get the standard curve of the relationship between color development distance and AHL concentration, which can be calculated by measuring the color development distance of the sa...

Embodiment 2

[0071] Embodiment 2, the application of AHL reporter strain in detecting the AHL concentration of Pseudomonas syringae tobacco pathogenic variety P.syringaepv.tabaci11528 (Pta) growth process in different growth stages

[0072] 1. Extract AHL

[0073] The AHL in the Pta culture medium in different growth phases (logarithmic phase, early stationary phase and late stationary phase) were extracted respectively. Concrete steps are as follows: add equal volumes of acidified ethyl acetate (containing 0.2% glacial acetic acid) to the Pta culture solution of different growth stages (logarithmic phase, early stable phase and post stable phase) respectively and extract three times; Phase (upper layer), combine the organic phase; blow dry with nitrogen, dissolve with chromatographic grade acetonitrile (honeywell, product number AH015-4); filter with organic filter membrane to obtain AHL extracts at different growth stages, and store them at -20°C for future use .

[0074] 2. Measure th...

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Abstract

The invention discloses a method for measuring the content of total acyl homoserine lactone (AHL). The method is established on the characteristic that an AHL biological report bacterial strain namely Agrobacterium tumefaciens A136 becomes blue when meeting AHL, and the color developing distance and the AHL concentration are positively correlated. The bacterial strain is used to detect whether a sample contains AHL or not, and can also be used to measure the AHL concentration. The AHL report bacterial strain (Agrobacterium tumefaciens A136) can be used to detect broad-spectrum AHLs; the sensitivities of the bacterial strain on AHL, whose acyl side chain length is C6-C14, are different; and the detection method based on the bacterial strain has the advantages of simpleness, rapidness, intuition, and precision, and can be widely applied to AHL detection and AHL concentration measurement.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for measuring the content of total acyl homoserine lactone (AHL) and an application thereof. Background technique [0002] Microbial cells can regulate gene expression in response to increasing cell density through quorum sensing (QS). Among them, QS means that as the density of microbial cells increases, the concentration of secondary metabolites produced by microbial cells also increases. When the concentration of a certain metabolite (also called a signal molecule) reaches a threshold value, microorganisms start to regulate various physiological functions. Related gene expression, including the formation of biofilm, the production of exopolysaccharide, the synthesis of virulence factors and antibiotics, etc. QS is a way of interaction and communication between cells that depends on the concentration of signal molecules that depend on cell density, and plays a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/02C12R1/01
CPCC12Q1/02
Inventor 庄国强程菲菲马安周
Owner RES CENT FOR ECO ENVIRONMENTAL SCI THE CHINESE ACAD OF SCI
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