GenoScreen's MIRU-VNTR typing solution is used by many healthcare centers, CNR and research teams worldwide, both as kits and as service.
MIRU-VNTR, a global reference
The MIRU-VNTR typing
The MIRU-VNTR (Mycobacterial Identification Repetitiv Unit-VNTR) analysis is an MLVA (Multiple Loci VNTR Analysis) analysis scheme specific to Mycobacterium tuberculosis. This is used in the genotyping of strains by numbering copies of VNTR present in 24 identified loci. The targeted VNTR loci are amplified by PCR with a set of primers specific for the flanking regions of each locus. The fragments thus obtained are processed by capillary electrophoresis. This method is the world standard for Mycobacterium tuberculosis typing, adopted by American and European CDCs and reference centers in more than 30 countries.
GenoScreen's MIRU-VNTR solution became the international standard reference for the MIRU-VNTR method (tests performed by RIVM 3, 4). It comes in a range of exclusive kits adapted to each step of the MIRU analysis (Calibration, validation and typing). This complete solution is optimized to deliver the best possible results with a standardized approach, while maintaining high reproducibility.
Learn more about our MIRU-VNTR kits
MIRU-VNTR Hypervariable, a scheme adapted to the strains of the Beijing lineage
M. tuberculosis strains identified as belonging to the so-called "Beijing" lineage cannot be distinguished by using the standard 24 loci. The sub-lineages in this family are identified by a MIRU analysis using a set of 4 additional hypervariables VNTR loci 1, 2. The MIRU-VNTR Hypervariable kit from GenoScreen enables this additional targeted analysis to be carried out.
GenoScreen's MIRU-VNTR solution is also available in a range specially designed for this complementary targeted analysis. The MIRU-VNTR Hypervariable kits (calibration, validation and typing) allow this specific typing, while benefiting from the same results quality and reproducibility as our MIRU-VNTR solution.
Learn more about our MIRU-VNTR Hypervariable kits
GenoScreen's expertise
With its cutting-edge technological expertise, GenoScreen is currently the leader worldwide in the genotyping and tracing of the Mycobacterium tuberculosis complex strains, the agent responsible for tuberculosis.
As kit or service packages, our solutions can be adapted to all issues:
- Research: structure of the strain population, evolution, comparison of virulence properties
- Public health: tuberculosis control and epidemiological surveillance
- Clinical trials: distinction between relapse and exogenous infection in case of treatment failure
- Clinical management: detection of cross-contamination, discrimination of close-related strains (clonal complexity)
The MIRU range, a complete offer
We propose kits and training for the implementation and use of the MIRU-VNTR method on Applied Biosystems® sequencers.
| MIRU-VNTR | Calibration kit | 1 calibration of Applied Biosystems® capillary sequencer and GeneMapper® software |
|---|---|---|
| Validation kit | 1 validation of the Applied Biosystems® capillary sequencer and GeneMapper® software settings | |
| Typing kit 24 markers according to international standard | 100 reactions (PCR + migration on Applied Biosystems® capillary sequencer) | |
| MIRU-VNTR hypervariable | Calibration kit | 1 calibration of Applied Biosystems® capillary sequencer and GeneMapper® software |
| Validation kit | 1 validation of the Applied Biosystems® capillary sequencer and GeneMapper® software settings | |
| Second line typing kit (4 markers) adapted to Beijing line strains | 50 reactions (PCR + migration on Applied Biosystems® capillary sequencer) | |
| MIRU-VNTR typing as service | Standard or customized schematics to meet your requirements | From DNA extracted (50 ng) or 50 µL of thermolysat on Applied Biosystems® capillary sequencer |
Calibration kit
Calibration of the analysis software and capillary sequencer is an essential preliminary step in the use of the GenoScreen MIRU-VNTR kit. We recommend a calibration of each sequencer and at each capillary change.
The MIRU-VNTR calibration kit contains: 14 reference DNA, 1 positive control (BCG), 6 master mixes (for the 24 VNTR loci), and an allelic ladder.
It provides calibration of one Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
The validation kit
The validation kit is used to validate the sequencer and software calibration over time. It is part of the quality control of the equipment.
The MIRU-VNTR validation kit contains: 14 reference DNA, 1 positive control (BCG) and 6 master mixes (for the 24 VNTR loci).
It provides the ability to validate the calibration of one Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
The 24 loci typing kit
The MIRU-VNTR typing kits contain the reagents necessary to perform MIRU typing on 24 VNTR loci, in accordance with the international standard. This kit allows analysis with a small amount of DNA and ensures a high rate of reproducibility of results.
The kit contains: a positive control (BCG) and 6 master mixes allowing the amplification of the 24 targeted VNTR loci.
It allows to run 100 tests on a previously calibrated Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
List of VNTR loci targeted by MIRU-VNTR 24-loci typing
| Loci | Alias | Loci | Alias | Loci | Alias | Loci | Alias |
|---|---|---|---|---|---|---|---|
| 154 | MIRU02 | 1644 | MIRU16 | 2401 | Mtub30 | 3171 | Mtub34 |
| 424 | Mtub04 | 1955 | Mtub21 | 2461 | ETRB | 3192 | Mtub34 |
| 577 | ETRC | 2059 | MIRU20 | 2531 | MIRU23 | 3690 | Mtub39 |
| 580 | MIRU04 | 2163b | QUB11b | 2687 | MIRU24 | 4052 | QUB26 |
| 802 | MIRU40 | 2165 | ETRA | 2996 | MIRU26 | 4156 | QUB4156 |
| 960 | MIRU10 | 2347 | Mtub29 | 3007 | MIRU27 | 4348 | MIRU39 |
The hypervariable calibration kit
Calibration of the analysis software and capillary sequencer is an essential preliminary step in the use of the GenoScreen MIRU-VNTR Hypervariable kit. We recommend a calibration of each sequencer and at each capillary change.
The MIRU-VNTR Hypervariable calibration kit contains: 6 reference DNA, 1 positive control (BCG), 1 master mix (for the 4 loci), and an allelic scale (ladder).
It provides calibration of one Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
The hypervariable validation kit
The validation kit allows to validate the sequencer and software calibration over time. It is part of the quality control of the equipment.
The MIRU-VNTR Hypervariable validation kit contains: 6 reference DNA, 1 positive control (BCG), and 1 master mix (for the 4 loci).
It provides the ability to validate the calibration of an Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
The hypervariable typing kit
The hypervariable MIRU-VNTR typing kit contains the reagents required to achieve MIRU typing on the additional 4 VNTR loci for differentiating Beijing sublines from each other. This kit allows accurate analysis with a small amount of DNA and ensures a high rate of reproducibility.
The kit contains: a positive control (BCG) and the master mix allowing the amplification of the 4 targeted VNTR loci.
It allows to run 50 tests on a previously calibrated Applied Biosystems® 3730, 3500, 3100 or 3130 capillary sequencer.
List of VNTR loci targeted by MIRU-VNTR Hypervariable typing 3, 4 :
| Loci | 1982 | 3820 | 3232 | 4120 |
|---|
Service provision
We also perform standard and customized MIRU-VNTR analyses as a service provider. We perform MIRU analyses on 9, 12, 15 or 24 standard VNTR markers and/or on the 4 loci dedicated to the Beijing lineage.
Available from 50 ng of extracted DNA (5 μL to 10 ng/µL) or 50 µL of thermolysate, our services include:
- PCR(s) realization ( according to the selected markers)
- PCR product migration on capillary sequencer
- Data analysis on GeneMapper
- Lineage assignment using the MIRU-VNTRplus database
Our training courses
GenoScreen created a complete training program dedicated to the MIRU-VNTR method. From the methodology to the results analysis, including the kits use and the sequencer calibration, these trainings will enable you to easily implement and/or perform the MIRU-VNTR analysis on a routine basis in your laboratory.
MIRU-VNTR training course :
- Theoretical aspects: the premise of the MIRU-VNTR typing method
- Practical aspects: training in the MIRU-VNTR kits use
- Implementation: training in the Applied Biosystems® sequencer calibration
- Data import & analysis: training in the GeneMapper® software use
- Data mining: training in the MIRU-VNTRplus analysis tool use
Références
1 Allix-Béguec C, Wahl C, Hanekom M, Nikolayevskyy V, Drobniewski F, Maeda S, Campos-Herrero I, Mokrousov I, Niemann S, Kontsevaya I, Rastogi N, Samper S, Sng LH, Warren RM, Supply P. Proposal of a consensus set of hypervariable mycobacterial interspersed repetitive-unit-variable-number tandem-repeat loci for subtyping of Mycobacterium tuberculosis Beijing isolates. J Clin Microbiol. 2014 Jan;52(1):164-72.
2 Trovato A, Tafaj S, Battaglia S, Alagna R, Bardhi D, Kapisyzi P, Bala S, Haldeda M, Borroni E, Hafizi H, Cirillo DM. Implementation of a Consensus Set of Hypervariable Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat Loci in Mycobacterium tuberculosis. Molecular Epidemiology. J Clin Microbiol. 2016 Feb;54(2):478-82.
3 de Beer JL, Kremer K, Ködmön C, Supply P, van Soolingen D; Global Network for the Molecular Surveillance of Tuberculosis 2009. First worldwide proficiency study on variable-number tandem-repeat typing of Mycobacterium tuberculosis complex strains. J Clin Microbiol., 2012 Mar;50(3):662-9
4 de Beer JL, Ködmön C, van Ingen J, Supply P, van Soolingen D; Global Network for Molecular Surveillance of Tuberculosis 2010. Second worldwide proficiency study on variable number of tandem repeats typing of Mycobacterium tuberculosis complex. Int J Tuberc Lung Dis., 2014 May;18(5):594-600