Primer composition for detecting drug resistance genes of mycobacterium tuberculosis drugs and application of primer composition

A technology of mycobacterium tuberculosis and primer composition, which is applied in the field of gene detection, can solve the problems of unreached and achieve the effects of reducing the missed detection rate, short detection cycle and high sequence coverage

Pending Publication Date: 2021-12-21
AGRI GENOMICS INST CHINESE ACADEMY OF AGRI SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0014] Although the above-mentioned patents have carried out sputum sample processing, identification of Mycobacterium tuberculosis and detection of drug resistance gene loci of some drugs, none of them have reached the level of replacing traditional drug sensitivity tests

Method used

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  • Primer composition for detecting drug resistance genes of mycobacterium tuberculosis drugs and application of primer composition
  • Primer composition for detecting drug resistance genes of mycobacterium tuberculosis drugs and application of primer composition
  • Primer composition for detecting drug resistance genes of mycobacterium tuberculosis drugs and application of primer composition

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0148] Example 1 A primer composition and kit for the detection of Mycobacterium tuberculosis drug resistance genes

[0149] 1. The primer composition is shown in Table 2 below.

[0150] Table 2 Primer Composition

[0151]

[0152]

[0153]

[0154]

[0155]

[0156]

[0157]

[0158]

[0159]

[0160]

[0161]

[0162] The drugs related to Mycobacterium tuberculosis include rifampicin (RIF), isonicotiny hydrazide (INH), ethambutol (ethambutol, EMB), streptomycin (Sm), Moxiza Star (moxifloxacin, MFX), levofloxacin (Levofloxacin, LFX), amikacin (AK), kanamycin (Kanamycin, Km), capreomycin (caperomycin, Cm), pyrazinamide (pyrazinamide, PZA ), rifabutin (RFB), ethionamide (Eto), clofazimin (Cfz), paraaminosalicylic acid (PAS), prothionamide (protionamide, PTO), cycloserine (cycloserine, Cs), bedaquiline (bedaquiline, Bdq), linezolid (Linezolid), Delamanid (Delamanid), Pretomanid (Pretomanid).

[0163] Specifically, the primer composition is use...

Embodiment 2

[0272] Embodiment 2 A kind of detection method of drug resistance gene of Mycobacterium tuberculosis

[0273] 1. Reagents

[0274] (1) DNA extraction kit: Purchased from QIAGEN Human Tissue and Cell DNA Extraction Kit, Cat. No. 51304.

[0275] (2) IGT-EM808 polymerase mixture, Enhancer buffer NB (1N), Enhancer buffer M, IGT-I5 Index (10 μM), IGT-I7 Index (10 μM): purchased from Aiji Taikang Biotechnology Company.

[0276] (3) Agencourt AMPure XP kit: including magnetic beads and YF buffer B, purchased from Beckman.

[0277] (4) DynaMag Side magnetic stand: purchased from Life Technologies.

[0278] 2. Experimental steps:

[0279] 2.1. Sample DNA extraction:

[0280] (1) Collect sputum and obtain samples.

[0281] (2) Using the DNA extraction kit, extract the total DNA from the sample, detect the DNA concentration, and obtain a qualified DNA extraction solution (the qualified DNA content requirement is >=50pg, and the bands are not degraded).

[0282] 2.2. Multiple amplif...

Embodiment 3

[0325] Example 3 Construction of Drug Resistance Effect Evaluation System

[0326] 1. Sample source:

[0327] (1) Retrospective samples: Whole-genome sequencing data and phenotypic drug susceptibility information of 19,673 Mycobacterium tuberculosis cultures in the PATRIC database;

[0328] (2) Confirmatory sample 1: Whole-genome sequencing data of 1961 Mycobacterium tuberculosis cultures and 9 kinds of phenotypic drug sensitivity information in the article "Rapid antibiotic-resistance predictions from genomesequence data for Staphylococcus aureus and Mycobacterium tuberculosis".

[0329] (3) Confirmatory sample 2: Whole 110 Mycobacterium tuberculosis cultures in the article "Evaluation of Whole-Genome Sequence Method to Diagnose Resistance of 13 Anti-tuberculosis Drugs and Characterize Resistance Genes in Clinical Multi-Drug Resistance Mycobacterium tuberculosis Isolates From China" Genome sequencing data and drug sensitivity information of 13 phenotypes.

[0330] 2. Formul...

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Abstract

The invention provides a primer composition and kit for detecting drug resistance genes of mycobacterium tuberculosis drugs, and belongs to the technical field of gene detection. The primer composition or the kit can be used for detecting 1200 sequence variation sites in 58 drug resistance genes related to 20 drugs for clinically treating mycobacterium tuberculosis, mycobacterium tuberculosis identification can be directly carried out on clinical extracts, culture is not needed, the detection number and detection coverage of the sequence variation sites are increased, the omission ratio of sequence variation sites is reduced, the detection specificity and sensitivity are high high, large-batch samples can be detected in parallel, and the detection cost is reduced. The primer composition and the kit thereof can be widely applied to scientific research, drug curative effect research, tuberculosis epidemic disease investigation and clinical medication guidance of tuberculosis.

Description

technical field [0001] The invention belongs to the technical field of gene detection, and in particular relates to a primer composition for detecting drug-resistant genes of Mycobacterium tuberculosis and its application in guiding the detection of drug-resistant tuberculosis drugs. Background technique [0002] Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis infection. Mycobacterium tuberculosis can invade various organs of the human body, 80% of which occur in the lungs, which is called tuberculosis, and other parts such as cervical lymph, meninges, peritoneum, intestines, skin, bones, etc. can also be secondary infected. Mycobacterium tuberculosis (Mycobacteriaceae tuberculosis), commonly known as tubercle bacillus (tubercle bacillus), is the pathogen that causes tuberculosis, which is a type of obligate aerobic bacteria, acid-fast staining positive, no flagella, pili, and microcapsules But it does not form spores, and its bacterial wal...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6858C12Q1/04C12N15/11
CPCC12Q1/689C12Q1/6858C12Q2600/106C12Q2600/156C12Q2600/16C12Q2531/113C12Q2537/143C12Q2535/122
Inventor 刘万飞曲久鑫林强崔鹏
Owner AGRI GENOMICS INST CHINESE ACADEMY OF AGRI SCI
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