Liquid-chip detection kit for multiple drug-resistant genes of pseudomonas aeruginosa

A Pseudomonas aeruginosa, detection kit technology, applied in the direction of determination/inspection of microorganisms, biochemical equipment and methods, etc., can solve the problems of long detection cycle, low detection throughput, and many influencing factors, and achieve specificity High, high detection throughput, accurate detection effect

Inactive Publication Date: 2015-01-28
中国人民解放军济南军区第四〇一医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, clinical laboratories mostly use drug-resistant phenotype detection methods. The main problems are long detection cycle, poor timeliness, low detection throughput, and many influencing factors, which cannot meet the current requirements of clinical anti-infection treatment and hospital infection control.

Method used

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  • Liquid-chip detection kit for multiple drug-resistant genes of pseudomonas aeruginosa
  • Liquid-chip detection kit for multiple drug-resistant genes of pseudomonas aeruginosa
  • Liquid-chip detection kit for multiple drug-resistant genes of pseudomonas aeruginosa

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Pseudomonas aeruginosa multi-drug resistance gene liquid chip detection kit, the Pseudomonas aeruginosa multi-drug resistance gene includes OPRD2, TEM, OXA10, AAC3, AAC6, VIM, VEB, IMP a total of 8 drug resistance genes, which It is characterized in that the kit includes nucleic acid amplification components and hybridization components;

[0027] The nucleic acid amplification components specifically include the following components:

[0028] Premix: Tris-HCl buffer containing dATP, dTTP, dCTP, dGTP, MgCl2;

[0029] Primer mixture: 8 pairs of primer mixtures for drug-resistant genes;

[0030] Polymerase: DNA polymerase containing glycerol and stabilizer formula; Positive quality control: positive strain mixture containing OPRD2, TEM, OXA10, AAC3, AAC6, VIM, VEB, IMP genes;

[0031] Negative quality control: sterile water;

[0032] Hybrid components specifically include the following components:

[0033] Microsphere hybridization solution: mixed suspension of microsp...

Embodiment 2 Embodiment 1

[0040] Specific verification of primers and probes in Example 2 Example 1:

[0041] 1. Take OPRD2, TEM, OXA10, AAC3, AAC6, VIM, VEB, and IMP-resistant Pseudomonas aeruginosa culture fluid respectively, and extract DNA. Prepare a multiplex PCR reaction system (40ul) using the above primers:

[0042]

[0043] 2. The prepared reaction system is amplified according to the following procedure:

[0044] a) The first stage: 95°C, 10min, 1 cycle.

[0045] b) Second stage: 95°C, 30sec; 56°C, 30sec; 72°C, 30sec; 40 cycles.

[0046] c) The third stage: 72°C, 10 minutes, 1 cycle.

[0047] 3. The amplified PCR product is analyzed by Luminex flow analyzer, the steps are as follows:

[0048] a. According to the number of tubes in the PCR reaction, use scissors to cut a microwell hybridization plate of the corresponding size. Turn on the Luminex instrument to preheat, and set the temperature of the microplate adapter copper plate on the instrument at 48°C.

[0049] b. Place the reage...

Embodiment 4

[0063] Embodiment 4 kit application verification

[0064] A certain number of clinical specimens were tested and analyzed using the kit in Example 1, and the specific results are shown in the table below. From the test results, 18 of the 40 samples were positive for drug-resistant genes, and 9 (50%) of them contained multiple drug-resistant genes.

[0065] The specimens come from the 401st Hospital of the People's Liberation Army, Qingdao Municipal Hospital, Qingdao University Affiliated Hospital, and various clinical departments of Qingdao Central Hospital. The types of specimens are mainly sputum and wound secretions. , Throat swab, bile.

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Abstract

The invention discloses a liquid-chip detection kit for multiple drug-resistant genes of pseudomonas aeruginosa. The eight multiple drug-resistant genes of the pseudomonas aeruginosa are OPRD2, TEM, OXA10, AAC3, AAC6, VIM, VEB and IMP. The kit comprises a nucleic-acid amplification component and a hybridization component, wherein the nucleic-acid amplification component comprises the following specific components: premixed liquid, primer mixed liquid, polymerase, a positive quality control product and a negative quality control product; and the hybridization component comprises the following specific components: microsphere hybridization liquid, quality control microspheres and streptavidin-phycoerythrin. The liquid-chip detection kit disclosed by the invention has the advantages that the multiple drug-resistant genes of the pseudomonas aeruginosa can be detected simultaneously, and the drug resistance of the pseudomonas aeruginosa can be rapidly and accurately detected, so that the need of clinical treatment is met; and simultaneously, the kit has the beneficial effects of high specificity, combined screening, high detection flux and more convenience in promotion and application.

Description

technical field [0001] The invention relates to a liquid chip detection kit for multiple drug resistance genes of Pseudomonas aeruginosa, which belongs to the technical field of biological detection. Background technique [0002] In 2010, as the term "super bacteria" frequently appeared in the newspapers, multi-drug resistance of bacteria and the rational use of antibiotics came back to people's attention. Super bacteria is actually not a new thing. It is not the name of a bacterium, but the name of a type of bacteria. The commonality of this type of bacteria is that they have strong resistance to almost all antibiotics. They have always existed and have been abused by humans. Strong resistance to antibiotics has evolved. Bacterial drug resistance, especially multidrug resistance (MDR) has become a very serious medical and social problem. [0003] The mechanism of bacterial drug resistance is very complex. Under the pressure of antibiotic selection, the clinical situation ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04
CPCC12Q1/689C12Q2600/16
Inventor 梁冰孙青菊黄宁
Owner 中国人民解放军济南军区第四〇一医院
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