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Quick investigating kit for melon bacterial fruit spot

A detection kit and bacterial technology, which is applied in the field of rapid detection kits for melon bacterial fruit rot, can solve the problems of no report on the identification of plant pathogenic bacteria, and achieve improved specificity and sensitivity, accurate detection, and sensitivity high effect

Inactive Publication Date: 2007-12-19
XINJIANG XIYU IND GROUP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Adsorption media are widely used in the fields of biochemistry and molecular biology to separate and purify biomolecules such as nucleic acids and proteins, but there is no report on the identification of plant pathogens

Method used

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  • Quick investigating kit for melon bacterial fruit spot

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1: In the embodiment of the present invention, the kit consists of two parts, namely:

[0037] 1. All supporting reagents and equipment for IAS-PCR detection of BFB.

[0038] 2. The technical operation and judgment standard of IAS-PCR detection.

[0039] Among them: 1. All supporting reagents for IAS-PCR detection of BFB are:

[0040] (1). A solution: BFB immunomagnetic beads 0.5mL

[0041] (2).B solution: PH6.8, 0.01mol / L phosphate PB buffer 10mL

[0042] (3).C solution: deionized water 1mL

[0043] (4).D solution: 10×PCR buffer, such as: MgCl 2 25mmol / L 0.24mL

[0044] (5).E solution: 2.5mmol / L, deoxynucleotide dNTP 0.006mL

[0045] (6).F solution: 0.5mmol / L primer BFB 1 0.006mL

[0046] (7).G solution: 0.5mmol / L primer BFB 2 0.006mL

[0047] (8).H solution: 2u / μTag enzyme 0.006mL

[0048] Among them, the detection BFB primer sequence:

[0049] BFB 1 : 5′-GAC CAG CCA CAC TGG GAC-3′

[0050] BFB 2 : 5′-C...

Embodiment 2

[0071] Embodiment 2: the preparation method of BFB immunomagnetic beads IAS in embodiment 1 is:

[0072] 1. Cultivate the standard BFB pathogen and collect the bacteria, extract the protein of the bacteria by ultrasonic crushing, make an adjuvant BFB antigen, immunize rabbits, and wait until the titer reaches 1:64, bloodletting to obtain anti-BFB serum, and use ammonium sulfate Analysis, FpLc protease A column purification to obtain BFB antibody IgG.

[0073] 2. Take 5 grams of solid carrier Sephadex or Cellulose with anion exchange groups, add 250 ml of sterile water, and let it swell overnight.

[0074] 3. Put 1ml of the above gel into a small chromatography column, elute 10ml with pH6.8, 0.01mol / L TB containing 0.4mol / L NaCl, and then use pH6.8, 0.01mol / L TB solution to balance wash Remove 20ml.

[0075] 4. Take 0.5ml of the purified BFB antibody containing 5mg of protein and add it to the above-mentioned small chromatography column. After the BFB antibody enters the colu...

Embodiment 3

[0076] Embodiment 3: In the embodiment of the present invention, the kit consists of two parts, namely:

[0077] 1. All supporting reagents and equipment for IAS-PCR detection of BFB.

[0078] 2. The technical operation and judgment standard of IAS-PCR detection.

[0079] Among them: 1. All supporting reagents for IAS-PCR detection of BFB are:

[0080] (1). A solution: BFB immunomagnetic beads 0.5mL

[0081] (2).B solution: PH6.8, 0.01mol / L phosphate PB buffer 10mL

[0082] (3).C solution: deionized water 1mL

[0083] (4).D solution: 10×PCR buffer, such as: MgCl 2 25mmol / L 0.24mL

[0084] (5).E solution: 2.5mmol / L, deoxynucleotide dNTP 0.006mL

[0085] (6).F solution: BFB primer and Tag enzyme 0.24mL

[0086] The detection technology operation of the kit is as follows:

[0087] (1). Sample treatment: Take 1000 seeds to be tested, add 200mL pH6.8 0.01mol / L PB buffer solution, shake gently on the shaker at room temperature for 4 hours, filter the seeds with 3 layers of gau...

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Abstract

A kit for quickly detecting bacterial corrosion disease of melon vegetables consists of main reagents as BFB immune magnetic beads, deionized water, phosphoric acid PB buffer liquid, PCR buffer liquid, deoxyribonucleotide dNTP and BFE primer or Tag enzyme. Its operating method and judgment standard are also disclosed.

Description

1. Technical field [0001] The invention belongs to a detection technology for the detection of melon bacterial fruit rot disease BFB caused by oat acidophilus subspecies watermelon, in particular to a rapid detection kit for melon bacterial fruit rot. 2. Background technology [0002] Acidovorax avenae subsp.citrulli, or Aac for short, can cause Bacterial fruit blotch of cucurbits, or BFB for short, which is a worldwide quarantine pest. In particular, the above-mentioned diseases are the three types of dangerous diseases stipulated in the "List of Dangerous Diseases, Insects and Weeds for Imported Plant Quarantine of the People's Republic of my country" published by our country. [0003] The incidence area of ​​melon bacterial fruit rot is expanding. The symptoms at the seedling stage of the disease often manifest as large black lesions without any water-soaked shape, which makes field diagnosis very difficult. The isolation and culture diagnosis of pathogenic bacteria takes...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N21/25G01N1/28G01N33/53C12Q1/00
Inventor 喻梅辉张红马光玉王叶筠周志成丁建军穆全昌姜明张兴平
Owner XINJIANG XIYU IND GROUP
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