Primer for RPA rapid test of salmonella carrying sulfonamides resistance gene and application of primer

A Salmonella and drug-resistant gene technology, applied in the field of RPA rapid detection primers, can solve the problems of unsatisfactory operation procedures and detection time specificity detection, and achieve good intra-species conservation and good inter-species specificity

Active Publication Date: 2017-06-27
ZHEJIANG GONGSHANG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is not ideal in terms of operating p...

Method used

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  • Primer for RPA rapid test of salmonella carrying sulfonamides resistance gene and application of primer
  • Primer for RPA rapid test of salmonella carrying sulfonamides resistance gene and application of primer

Examples

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Embodiment 1

[0028] Step (1). Sample DNA extraction

[0029] Take 1ml of the cultured bacteria solution and add it to a 1.5ml centrifuge tube, centrifuge at 12000g for 1min, discard the supernatant, add 400μl lysate to the centrifuge tube, mix well, add 600μl phenol-chloroform-isoamyl alcohol mixed solvent, shake vigorously, and centrifuge at 12000g 10min; take the supernatant, add chloroform-isoamyl alcohol mixed solvent equal to the volume of the supernatant, shake vigorously, and centrifuge at 12000g for 10min; take the supernatant, add chloroform equal to the volume of the supernatant, shake vigorously, Centrifuge at 12,000 g for 10 min; take the supernatant, add isopropanol 0.8 times the volume of the supernatant, and centrifuge at 12,000 g for 10 min; take the precipitate, wash it once with ethanol with a volume content of 70%, and then dry it at room temperature or 50°C Dry for 20 minutes, then add 80 μl of TE buffer solution with a pH value of 8.0 to dissolve to obtain a DNA extrac...

Embodiment 2

[0042] Step (1). Sample DNA extraction

[0043] Take 1ml of the cultured bacteria solution and add it to a 1.5ml centrifuge tube, centrifuge at 12000g for 1min, discard the supernatant, add 400μl lysate to the centrifuge tube, mix well, add 600μl phenol-chloroform-isoamyl alcohol mixed solvent, shake vigorously, and centrifuge at 12000g 10min; take the supernatant, add chloroform-isoamyl alcohol mixed solvent equal to the volume of the supernatant, shake vigorously, and centrifuge at 12000g for 10min; take the supernatant, add chloroform equal to the volume of the supernatant, shake vigorously, Centrifuge at 12,000 g for 10 min; take the supernatant, add isopropanol 0.8 times the volume of the supernatant, and centrifuge at 12,000 g for 10 min; take the precipitate, wash it once with ethanol with a volume content of 70%, and then dry it at room temperature or 50°C Dry for 20 minutes, then add 80 μl of TE buffer solution with a pH value of 8.0 to dissolve to obtain a DNA extrac...

Embodiment 3

[0056] Step (1). Sample DNA extraction

[0057] Take 1ml of the cultured bacteria solution and add it to a 1.5ml centrifuge tube, centrifuge at 12000g for 1min, discard the supernatant, add 400μl lysate to the centrifuge tube, mix well, add 600μl phenol-chloroform-isoamyl alcohol mixed solvent, shake vigorously, and centrifuge at 12000g 10min; take the supernatant, add chloroform-isoamyl alcohol mixed solvent equal to the volume of the supernatant, shake vigorously, and centrifuge at 12000g for 10min; take the supernatant, add chloroform equal to the volume of the supernatant, shake vigorously, Centrifuge at 12,000 g for 10 min; take the supernatant, add isopropanol 0.8 times the volume of the supernatant, and centrifuge at 12,000 g for 10 min; take the precipitate, wash it once with ethanol with a volume content of 70%, and then dry it at room temperature or 50°C Dry for 20 minutes, then add 80 μl of TE buffer solution with a pH value of 8.0 to dissolve to obtain a DNA extrac...

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Abstract

The invention discloses a primer for RPA rapid test of salmonella carrying a sulfonamides resistance gene and application of the primer. The primer has better interspecies specificity and intraspecies conservativeness, the technology can detect salmonella and sulfonamides resistance gene (sul3) in one reaction, whether the bacterium is the salmonella carrying the sulfonamides resistance gene (sul3) can be judged in short time, no false positive occurs, expensive instruments are not needed, and accordingly, the primer is more convenient, efficient and accurate. The segments after amplification are different is length and can be distinguished by electrophoresis.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a primer for rapid detection of Salmonella and sulfa drug-resistant genes (Recombinase Polymerase Amplification, RPA) primers and applications thereof, in particular to a method capable of simultaneously detecting Salmonella and sulfonamides. Primers for rapid detection of RPA of sulfa drug resistance gene (sul3) and its application. Background technique [0002] Salmonella (Salmonella) is a common zoonotic pathogen in Enterobacteriaceae. It can not only cause pullorum, piglet paratyphoid, abortion and other animal diseases, but also cause typhoid fever, paratyphoid fever, sepsis, gastroenteritis and food poisoning in humans , in food poisoning around the world, the poisoning cases caused by Salmonella account for the first or second place. Sulfa drugs have the advantages of low price, easy storage, broad antibacterial spectrum, and remarkable efficacy, and are still necessary drugs f...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/10C12N15/11C12R1/42
CPCC12Q1/6844C12Q1/689C12Q2600/106C12Q2521/507C12Q2521/101C12Q2565/125Y02A50/30
Inventor 曲道峰韩剑众项紫莹沈杨朱安怡齐艳君康震宋竹青魏杰
Owner ZHEJIANG GONGSHANG UNIVERSITY
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