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Method for detecting ten kinds of trimethoprim drug resistance genes and kits used

A technology of trimethoprim and drug-resistant genes, which is applied in the fields of biochemical equipment and methods, and the measurement/testing of microorganisms, etc., which can solve the problems of inability to detect, unknown gene sequence, inability to design specific primers and probes, etc. low cost, high sensitivity, and good specificity

Inactive Publication Date: 2020-05-05
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0014] Although the results of genotype detection are accurate, it can only detect drug-resistant genes with known sequences, and for new drug-resistant genes that have not been discovered, because the gene sequence is unknown, specific primers and probes cannot be designed in a targeted manner.
In contrast, phenotypic testing is still possible to obtain positive detection results for drug resistance caused by novel drug resistance genes because it directly detects the survival status of strains under drug selection pressure

Method used

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  • Method for detecting ten kinds of trimethoprim drug resistance genes and kits used
  • Method for detecting ten kinds of trimethoprim drug resistance genes and kits used
  • Method for detecting ten kinds of trimethoprim drug resistance genes and kits used

Examples

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Embodiment 1

[0046] Embodiment 1, using the present invention to detect trimethoprim drug-resistant genes

[0047] In this example, all trimethoprim drug resistance genes involved in the present invention were detected.

[0048] 1. Preparation of oligonucleotide primers and probes

[0049] The present invention designs 3-5 pairs of specific amplification primers for each involved trimethoprim drug-resistant gene according to the reference sequence provided by the CARD database, combined with the sequence retrieved in the Genbank database, and then passes The experiment screened out the best specific primer combination (sequence 1-sequence 20 as shown in the attached sequence table); according to the sequence specificity of each trimethoprim drug-resistant gene involved, for each A drug-resistant gene designed 1-5 different specific probes, and then screened through experiments to select specific probes that meet the requirements of sensitivity and specificity (sequence 21-sequence 30 as s...

Embodiment 2

[0081] Example 2: Using the present invention to detect trimethoprim drug resistance genes in clinical samples

[0082] This example illustrates that the method and kit provided by the present invention can be used for the detection of trimethoprim drug resistance genes in clinically isolated bacterial strains.

[0083] 1. Preparation of Oligonucleotide Primers and Probes

[0084] The oligonucleotide primers used in this embodiment are the same as in Example 1; the specific probes used are the specific probes (sequence 21-30) selected on the basis of Example 1, and the method and steps are the same as in Example 1 .

[0085] 2. Preparation of a chip for detection of trimethoprim drug resistance genes

[0086] The probe dot matrix arrangement method used in this embodiment is according to image 3 Shown, other methods and steps are with embodiment 1.

[0087] 3. Preparation of samples to be tested

[0088] Pseudomonas aeruginosa (sample number Pae117), Klebsiella pneumonia...

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Abstract

The invention discloses a kit for detecting drug resistance genes of ten trimethoprim drugs. The kit comprises an amplification primer combination for carrying out specific amplification on the drug resistance genes of the ten trimethoprim drugs and a specific probe for detecting on the drug resistance genes of the ten trimethoprim drugs. The invention further discloses a method for detecting thedrug resistance genes of the ten trimethoprim drugs by using the kit. The method for detecting the drug resistance genes of the trimethoprim drugs and the kit thereof have the characteristics of highintegration degree, high sensitivity, good specificity, stable and reliable detection results and low cost, can be widely used for distribution and epidemiological monitoring of the drug resistance genes of the trimethoprim drugs, and provides an effective tool for rational drug use and effective reduction and delay of drug resistance.

Description

technical field [0001] The invention relates to a method for detecting drug-resistant genes of 10 kinds of trimethoprim drugs and a kit thereof. Background technique [0002] 1. Trimethoprim drugs and their mechanism of action: [0003] Trimethoprim (TMP) is a class of synthetic antibacterial agents. The first TMP compound was first used in the UK in 1962. Since the combination of TMP-SUL showed a synergistic effect in vitro, since 1968, TMP has often been used in combination with sulfonamide (SUL) drugs. However, clinical experience has shown that the TMP-SUL combination does not have a significant synergistic effect in vivo. TMP has been used alone since the 1970s, first for the prevention of UTIs and then for the treatment of acute UTIs. [0004] TMP can bind to dihydrofolate reductase (DHFR; EC 1.5.1.3) in bacteria, inhibit dihydrofolate reductase, and hinder the reduction of dihydrofolate to tetrahydrofolate. Tetrahydrofolate is necessary for the synthesis of folat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12Q1/6834
CPCC12Q1/6834C12Q1/6883C12Q2600/106C12Q2600/16C12Q2563/107C12Q2531/107C12Q2537/143
Inventor 张琼李兰娟项春生刁宏燕刘芳孟晓华姚坚周佳维吴灵娇倪淑君
Owner ZHEJIANG UNIV
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