LAMP (Loop-mediated Isothermal Amplification) detection kit and method of pathogenic aeromonas hydrophila
The technology of Aeromonas hydrophila and detection kit is applied in the field of detection of target DNA fragments, and can solve the problems of numerous detection steps for pathogenic Aeromonas hydrophila, difficult to grasp the accuracy, time-consuming data processing and the like, To achieve the effect of reducing background influence, method convenience, and enhancing specificity
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[0052] The present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments.
[0053] 1) DNA extraction
[0054] Pick suspected colonies and add them to 0.4mL TE buffer (pH8.0) for shaking and mixing. Add 100 μL of lysozyme solution, incubate at 37°C for 10 minutes, add 10 μL of proteinase K solution, incubate at 55°C for 1-2 hours, add the same volume of Tris saturated phenol (pH8.0), shake vigorously, centrifuge at 11000g for 3 minutes, and take the supernatant solution, and repeated phenol extraction; take the supernatant, add 0.1 times the volume of sodium acetate (3mol / L), mix well, add an equal volume of ice ethanol, mix well, let stand at low temperature for 30 minutes, and centrifuge at 15000g for 5 minutes. Discard the supernatant, wash once with 70% cold ethanol, centrifuge at 15,000 g for 5 minutes at room temperature, discard the supernatant, add 50 μL TE solution to obtain a genomic DNA solution to be teste...
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