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A fluorescent probe that can be used to detect cutinase on the surface of plant leaves and its preparation method

A technology of fluorescent probes and plant leaves, which is applied in the field of analysis and testing, can solve the problems of high requirements for operation methods and cumbersome steps, and achieve the effects of improving detection efficiency, simple operation, and convenient results

Active Publication Date: 2021-08-24
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to overcome the shortcomings of cumbersome steps and high requirements for operating means in the field of plant disease detection, and use cutinase as a plant disease marker to provide a fluorescent probe that can be used to detect cutinase on the surface of plant leaves and its preparation method , the detection process is simple and fast, and the result judgment is intuitive, which is conducive to the mastery and application of this technology by non-technical personnel

Method used

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  • A fluorescent probe that can be used to detect cutinase on the surface of plant leaves and its preparation method
  • A fluorescent probe that can be used to detect cutinase on the surface of plant leaves and its preparation method
  • A fluorescent probe that can be used to detect cutinase on the surface of plant leaves and its preparation method

Examples

Experimental program
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Effect test

Embodiment 1

[0045] An embodiment of the fluorescent probe of the present invention and its preparation method, the fluorescent probe of this embodiment is N-n-butyl-4-(4-butyryloxystyryl)-1,8-naphthalene Diimide, its structural formula is as shown in (I):

[0046]

[0047] The synthetic route of this embodiment is as figure 1 Shown, its specific preparation method is as follows:

[0048] 1) Dissolve the reaction raw material 4-bromo-1,8-naphthalene diimide (0.5g, 1.8mmol) in 5ml ethanol and put it into a 25ml two-necked bottle and put a stirring bar in it, then place the two-necked bottle Stir on a magnetic stirrer for 15 minutes. After it is completely dissolved, use a pipette gun (100-1000μl) to accurately measure 0.36ml of n-butylamine (0.36ml, 3.6mmol) into the two-necked bottle, and then assemble the serpentine return tube into the On the two-necked bottle, and nitrogen protection was introduced through the other port of the two-necked bottle, and then the crystallization dish c...

Embodiment 2

[0052] The anti-moisture interference of embodiment 2 fluorescent probe

[0053] Add 10%, 20%, 40%, 60%, 70%, 80%, and 90% of the water in the system to N-n-butyl-4-(4-butyryloxystyryl)-1,8- The ability of anti-moisture interference in the detection process of the probe was observed in the ethanol solution of naphthalene diimide. The result is as Figure 4 As shown, the probe can complete the normal detection of cutinase when the environmental moisture content is less than 80%, and has high practical application potential.

Embodiment 3

[0054] Example 3 Ratiometric detection of cutinase in the environment by N-n-butyl-4-(4-butyryloxystyryl)-1,8-naphthalene diimide.

[0055] The fluorescent probe N-n-butyl-4-(4-butyryloxystyryl)-1,8-naphthalene diimide prepared in Example 1 is used for ratiometric detection of cutinase in the environment . Use a probe solution with a concentration of 4 μM during the test, add a fixed concentration of cutinase (70U / L) to the probe solution, record the fluorescence intensity of the solution at 520nm and 580nm after reacting for different times, and calculate based on this The detection ratio I580 / I520 is obtained to achieve the purpose of ratio detection. Such as Figure 5 As shown, the lines 1-8 in the figure represent the fluorescence images when the reaction time between the probe solution and cutinase is 0 min, 5 min, 10 min, 15 min, 20 min, 25 min, 30 min, 35 min, and 40 min, respectively. It can be seen from the figure that the fluorescence emission position of the prob...

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Abstract

The invention relates to a fluorescent probe capable of detecting cutinase on the surface of plant leaves and a preparation method thereof, belonging to the technical field of analysis and detection. The specific chemical structure of the fluorescent probe of the present invention is shown in formula (I). The fluorescent probe can detect the cutinase produced by phage on the surface of plant leaves with specificity and anti-interference, and the fluorescent color of the probe will also undergo a red shift visible to the naked eye after reacting with the cutinase, which can realize the detection of cutinase. Ratiometric detection of enzymes. Secondly, the probe can also be widely used in the detection of liquid environment, gas environment, and chemical species cutinase. At the same time, the probe has good stability, and has the characteristics of high sensitivity, high selectivity and specificity for the detection of cutinase.

Description

technical field [0001] The invention relates to a fluorescent probe that can be used for detecting cutinase on the surface of plant leaves and a preparation method thereof, belonging to the technical field of analysis and testing. Background technique [0002] Cutinase is a kind of serine esterase, which mainly acts on the hydrolysis of the ester bonds existing in plant cutin, and can also act on esters such as long-chain fatty acids and short-chain fatty acids. Cutinase can be produced by bacteria, fungi and artificially. Cutinase is a protein with a common structural framework of α / β hydrolytic folding, and the composition and structure of cutinase produced by different pathogens are basically the same. At the same time, cutinase is an inducible enzyme, and its main component is glycoprotein, which is generally combined with carbohydrates through O-glycosidic bonds. The current main application fields of cutinase are relatively narrow, mainly food industry and textile in...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07D221/14C09K11/06G01N21/64
CPCC07D221/14C09K11/06C09K2211/1007C09K2211/1029G01N21/643G01N2021/6417
Inventor 侯贤锋高振忠王晓波陈凯李锰
Owner SOUTH CHINA AGRI UNIV
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