Stenotrophomonassp. H-4 and preparation method of degrading enzyme preparation thereof as well as application

A Stenotrophomonas, unit technology, applied in the field of microbiology, can solve the problems of no report on the degradation effect of pesticide residues in fruits and vegetables, and the undiscovered germplasm resources of Stenotrophomonas.

Inactive Publication Date: 2013-09-18
上海阿斯沃特企业发展有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, germplasm resources of Stenotrophomonas that can degrade chlorothalonil have not been found
At present, there is no report on the degradation effect of Stenotrophomonas degrading enzyme preparation on pesticide residues in fruits and vegetables

Method used

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  • Stenotrophomonassp. H-4 and preparation method of degrading enzyme preparation thereof as well as application
  • Stenotrophomonassp. H-4 and preparation method of degrading enzyme preparation thereof as well as application
  • Stenotrophomonassp. H-4 and preparation method of degrading enzyme preparation thereof as well as application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Example 1: Screening identification and degradation characteristics of chlorothalonil-degrading bacteria

[0019] 1.1 Materials and methods

[0020] 1.1.1 Test materials

[0021] Basal salt medium (degradation medium, MM): 0.5g KH 2 PO 4 , 1.2 g Na 2 HPO 4 12H 2 O, 0.5g NH 4 NO 3 , 0.2g MgSO 4 ·7H 2 O, 50mg FeSO 4 , 10mL trace element solution (50mg Ca(OH) per liter 2 , 30 mg H 3 BO 3 , 10mg ZnSO 4 ·7H 2 O, 5 mg MnSO 4 ·H 2 O, 5 mg CuSO 4 ·5H 2 O, 5 mg Na 2 MoO 4 2H 2 O) Add ultrapure water to 1 L. Use 2mol / L HCl and 2mol / L NaOH solutions to adjust the pH value of the medium to 6.8-7.2. In order to avoid the influence of chloride ions in the water on the experimental process, the medium is prepared with ultrapure water. Add chlorothalonil to basal salt medium as the sole carbon source;

[0022] LB medium: peptone 10.0g, yeast powder 5.0g, NaCl 10.0g, add ultrapure water to 1L, adjust the pH value of the medium to 6.8-7.2 with 2mol / L HCl and 2mol / ...

Embodiment 2

[0051] Embodiment 2: the fermentation condition of chlorothalonil-degrading bacteria seed liquid and intracellular degradation enzyme extraction method optimization

[0052] 2.1 Materials and methods

[0053] 2.1.1 Test materials

[0054] Test strain: Stenotrophomonas (referred to as H-4), isolated and screened from red soil [preservation number: CGMCC No.7247]. First inoculate the strains on LB solid plate medium, cultivate and activate at 28°C, and then pick single colonies into LB liquid medium and cultivate for 24 hours.

[0055] LB medium (g / L): peptone 10.0, yeast powder 5, NaCl 10, deionized water 1L, pH7.0-7.2, sterilized at 121°C for 20min.

[0056] PBS phosphate buffer (g / L): NaCl 8, KCl 0.2, NaCl 2 HPO 4 12H 2 O 3.64, KH 2 PO 4 0.24, add ultrapure water to 1L, pH7.4.

[0057] 2.1.2 Orthogonal test of seed liquid fermentation

[0058] According to three factors and three levels, A: bacterial inoculation amount (A1=5%vt, A2=8%vt, A3=10%vt); B: culture temper...

Embodiment 3

[0072] Example 3: The cleaning effect of degrading enzyme preparations on chlorothalonil on different vegetables

[0073] 3.1 Materials and methods

[0074] 3.1.1 Test materials

[0075] Preparation of contaminated vegetables: Use 75% agricultural chlorothalonil powder, dissolve 0.5g of chlorothalonil powder in 10L of water, immerse cucumbers, tomatoes and green vegetables in it, soak for 30 minutes, take out the vegetables, rinse them lightly with water, and place them in ventilation Dry in the cupboard.

[0076] Preparation of degrading enzyme solution: using optimized culture conditions, expand the cultivation of Stenotrophomonas, centrifuge the cultured bacterial suspension at 10000r / min for 10min, wash the obtained precipitate twice with PBS phosphate buffer solution of pH 7.4, dissolve In 20mL PBS phosphate buffer solution. Under the condition of 4°C ice bath, the cells were broken with an ultrasonic instrument (ultrasonic power: 30%), the ultrasonic time was 5 min, a...

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Abstract

The invention provides stenotrophomonassp. H-4 and a preparation method of degrading enzyme preparation thereof as well as application. The stenotrophomonassp. H-4 has been conserved in the conservation unit specified by the State Intellectual Property Office; the conservation date is February 1, 2013; the name of the conservation unit is China General Microbiological Culture Collection Center; and the conservation number is CGMCC No.7274. The stenotrophomonassp. H-4 can be grown on a culture medium with chlorothalonil as the unique carbon source and energy, and after being cultivated for 7 days, the degradation rate of the stenotrophomonassp. H-4 reaches 83.6%; the enzyme activity is 3.28 U and the degradation rate of the degrading enzyme to the chlorothalonil is 86% when the extraction conditions for crude enzyme in the stenotrophomonassp. H-4 are as follows: ultrasonic powder: 30 W; and ultrasonic time: 5 minutes. Obviously, the degrading enzyme preparation provided by the invention has bright application prospect in clearing away residual pesticides on vegetables and fruits.

Description

technical field [0001] The invention belongs to the technical field of microorganisms, and in particular relates to a preparation method and application of a pesticide fungicide chlorothalonil-degrading bacterium Stenotrophomonas H-4 and a degrading enzyme preparation thereof. Background technique [0002] China is a big agricultural country and also a big pesticide consumer. Especially with the rapid development of facility agriculture, the high-frequency and high-dose input of a large number of pesticides has caused agricultural products and soil pollution, resulting in frequent occurrence of pesticide residue accidents in fruit and vegetable products, seriously affecting the fruits and vegetables of our country. Product quality and safety. Therefore, the quality and safety of fruits and vegetables has become an important factor restricting the development of my country's fruit and vegetable industry. Strengthening the quality and safety control of fruits and vegetables ha...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/00A23L1/015C12R1/01A23L5/20
Inventor 滕应张满云马文亭陈婷骆永明李振高赵其国陈勇姜志伟蒋锐
Owner 上海阿斯沃特企业发展有限公司
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