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Nucleic acid constant-temperature amplification method of Methicillin-resistant Staphylococcus aureus (MRSA)

A methicillin, staphylococcus-resistant technology, applied in the field of bacterial biological detection, can solve problems such as no research reports yet

Pending Publication Date: 2020-01-31
SHANGHAI RENDU BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the application of SAT technology in the detection of different types of viruses and bacteria faces different problems, and it is necessary to specifically analyze the characteristics of viruses and bacteria for special design.
At present, there is no research report on real-time fluorescent nucleic acid constant temperature amplification detection technology for methicillin-resistant Staphylococcus aureus (MRSA)

Method used

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  • Nucleic acid constant-temperature amplification method of Methicillin-resistant Staphylococcus aureus (MRSA)
  • Nucleic acid constant-temperature amplification method of Methicillin-resistant Staphylococcus aureus (MRSA)
  • Nucleic acid constant-temperature amplification method of Methicillin-resistant Staphylococcus aureus (MRSA)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0185] Example 1 Design of special primers and probes for real-time fluorescent nucleic acid constant temperature amplification detection of methicillin-resistant Staphylococcus aureus (MRSA)

[0186] The present invention selects highly conserved and specific segments in the MRSA (SA 23s rRNA and mecA) gene as the amplified target sequence region (the nucleotide sequences thereof are shown in SEQ ID No.: 1, SEQ ID No.: 2 ), according to the principle of primer probe design, using DNAStar, DNAMAN software and manual design for real-time fluorescent nucleic acid constant temperature amplification detection of methicillin-resistant Staphylococcus aureus (SA 23s rRNA and mecA of MRSA) special primers and probe sequences , to obtain the following specific sequence:

[0187] (1) A capture probe (TCO, Target Capture Oligo) that can specifically bind to the target nucleic acid (SA 23s rRNA) sequence of methicillin-resistant Staphylococcus aureus (MRSA), the nucleotide sequence of the...

Embodiment 2

[0214] Example 2 Preparation of a real-time fluorescent nucleic acid constant temperature amplification detection kit for methicillin-resistant Staphylococcus aureus (MRSA)

[0215] Using the special primers and probes provided in Example 1, the real-time fluorescent nucleic acid constant temperature amplification detection kit for methicillin-resistant Staphylococcus aureus (MRSA) of the present invention was obtained. The kit contains capture probe (TCO, TargetCapture Oligo), T7 primer, nT7 primer, MRSA (SA 23srRNA) detection probe, SA internal standard detection probe, MRSA (mecA) detection probe, mecA internal standard detection probe Needle, internal standard, M-MLV reverse transcriptase and T7 RNA polymerase and other components.

[0216] The capture probe is present in the nucleic acid extraction solution, and the T7 primer, nT7 primer, MRSA detection probe, and internal standard detection probe are respectively present in the MRSA (SA 23srRNA) detection solution and th...

Embodiment 3

[0244] Example 3 Methicillin-resistant Staphylococcus aureus real-time fluorescence nucleic acid constant temperature amplification detection method

[0245] 1. Sample collection, transportation and storage

[0246] sample collection

[0247] 1.1 Collection of nasal and oropharyngeal swab specimens: Take nasal and oropharyngeal secretions with medical cotton swabs, soak the swab head in 2mL of normal saline and stick to the tube wall to squeeze dry, add 2mL of sample preservation solution to it for later use, The sample is the sample to be tested.

[0248] 1.2 Wound secretion sample collection: Take the wound secretion with a medical cotton swab, soak the swab head in 2mL of normal saline and stick it to the wall of the tube to squeeze dry, add 2mL of sample preservation solution to it for later use, and this sample is the sample to be tested.

[0249] Specimen Transport and Storage

[0250] The collected specimens can be used for testing immediately or stored at -70°C for ...

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Abstract

The invention discloses a nucleic acid constant-temperature amplification method of Methicillin-resistant Staphylococcus aureus (MRSA). Specifically, the method disclosed by the invention comprises the following step: performing amplification in a reaction system, wherein the reaction system comprises two specific primer pairs of Methicillin-resistant Staphylococcus aureus (MRSA); and the primersare capable of amplifying amplification products corresponding to two characteristic sequences (SA 23s rRNA and mecA RNA) of the Methicillin-resistant Staphylococcus aureus (MRSA) from a detection sample with a very low MRSA copy number. The method disclosed by the invention is capable of amplifying a to-be-detected sample with MRSA (SA 23s rRNA and mecA RNA) with high specificity, high sensitivity, low pollution and high rapidness, has the characteristics of high detection efficiency and high accuracy and has wide application prospects.

Description

technical field [0001] The invention relates to the technical field of biological detection of bacteria, in particular to the use in the real-time fluorescent nucleic acid constant temperature amplification detection of methicillin-resistant Staphylococcus aureus (MRSA) which combines specific target capture technology and real-time fluorescent nucleic acid constant temperature amplification detection technology Primers, probes and related kits. Background technique [0002] Staphylococcus aureus (Staphylococcus aureus, SA, referred to as Staphylococcus aureus) is an important pathogenic bacteria in medicine. , MRSA) infection is multi-drug resistant and has a high mortality rate, which has become a difficult point in clinical anti-infection treatment. Staphylococcus aureus is the most common pathogenic bacteria in human suppurative infection, which can cause local suppurative infection, pneumonia, pseudomembranous enteritis, pericarditis, etc., and even serious systemic in...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/14C12N15/11
CPCC12Q1/689C12Q1/6844C12Q2600/166C12Q2531/119C12Q2563/107C12Q2545/101
Inventor 商学军居金良于明辉崔振玲
Owner SHANGHAI RENDU BIOTECH
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