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Aeromonas hydrophila aptamer and its screening method and application

A technology of Aeromonas hydrophila and aptamer, which is applied in the field of Aeromonas hydrophila aptamer and its screening, which can solve the problems of cumbersome detection process, high cost, and time-consuming

Inactive Publication Date: 2016-07-06
JIMEI UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

At present, the detection of Aeromonas hydrophila has always been a thorny problem. The traditional chemical pathological detection process is cumbersome and time-consuming; enzyme-linked immunosorbent immunoassay and multiplex PCR technology are too expensive (2. Wang Li. Fish Aeromonas hydrophila Several detection methods of bacteria [J]. Inland Fisheries, 2006, (2): 22-23.)

Method used

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  • Aeromonas hydrophila aptamer and its screening method and application
  • Aeromonas hydrophila aptamer and its screening method and application
  • Aeromonas hydrophila aptamer and its screening method and application

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Embodiment Construction

[0023] 1. Synthesis of ssDNA library and primers

[0024] Referring to the method disclosed in the literature (7. Liu Fengwei, Lan Xiaopeng. In vitro screening of Pseudomonas aeruginosa aptamer research and preliminary application [D]. Fuzhou Fujian Medical University, 2007.), the random ssDNA library used for SELEX screening and primers were synthesized by Shanghai Sangon Bioengineering Technology Service Co., Ltd.

[0025] The library used was: 5'-GGGAGCTCAGAATAAACGCTCAA-N35-TTCGACATGAGGCCCGGATC-3'.

[0026] The primers used are:

[0027] Primer Ⅰ: 5'-GGGAGCTCAGAATAAACGCTCAA-3';

[0028] Primer II: 5'-GATCCGGGCCTCATGTCGAA-3';

[0029] Primer III: 5′-Digoxigenin-GGGAGCTCAGAATAAACGCTCAA-3′;

[0030] Primer IV: 5'-Biotin-GATCCGGGCCTCATGTCGAA-3'.

[0031] After the above primers were synthesized, they were diluted with TE buffer (pH 8.0) commonly used in molecular biology to a concentration of 10 μmol, and stored at -20°C for future use.

[0032] 2. Experimental reagents a...

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Abstract

The present invention involves a moderate and screening methods and applications of aquatic monolia.Provide a screening method and application of water -adding monolithic body and the method of screening of water -only gas monolithic bodies.The specific steps of the screening method of water -enchanting monocyloburates include: to build and synthesize the SSDNA library of hydrophilic monolithium bacteria that is to be selected, and design the corresponding primer; take an appropriate amount of SSDNA library and addictionA combination of water and gas monocular bacteria; SSDNA, which is separated from the combination of water -based monolithium, is separated, and the SSDNA is used as a template for PCT amplification. The amplification product is screened again until the corresponding appropriate body.The present invention uses Selex technology to screen the high -affinity and oligonucleotide of the pathogenic monolithic bacteria, which can be widely used in the detection and analysis of hydrophobic bacteria.

Description

technical field [0001] The invention relates to common pathogenic bacteria in aquaculture—Aeromonas hydrophila, in particular to the Aeromonas hydrophila aptamer and its screening method and application. Background technique [0002] Aeromonas hydrophila (Aeromonas hydrophila) is widely distributed in nature and is the primary pathogenic bacteria of various aquatic animals. It can produce a variety of pathogenic factors and is pathogenic to aquatic animals, livestock and humans. It causes sepsis in various animals and human diarrhea, which poses a threat to human health and also causes large economic losses to the breeding industry (1. Zhang Cuijuan, Yu Zhouliang, Zhao Baohua, He Hongxuan. Research progress on Aeromonas hydrophila [J]. Chinese Journal of Veterinary Medicine, 2008,42(7):46-50.). Therefore, for the pollution of Aeromonas hydrophila, which is an indicator bacterium of water source pollution, the detection strength still needs to be further strengthened. At pr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/115C12Q1/68C12Q1/04C12R1/01
Inventor 李元跃王雷段博文陈融斌黎中宝
Owner JIMEI UNIV
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