Genotyping

GenoScreen proposes you the opportunity to characterize more accurately your microorganisms (subspecies or strain characterization) using standardized and validated molecular biology methods such as MLST or MLVA (including MIRU-VNTR).

MultiLocus Sequencing Typing (MLST)

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The MLST (MultiLocus Sequencing Typing) sequencing typing is the reference technique for the distinction of different strains. This method is based on the house-keeping genes sequencing (house-keeping genes are encoding essential proteins of the bacterium).

These sequences have the particularity of presenting a stable polymorphism over time and sufficient enough to distinguish strains from each other.

This approach allows the characterization of a microorganism whose genus (or species) is known in order to identity the species(or subspecies) and this thanks to the sequencing of 7 house-keeping genes.

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The MLST by GenoScreen

GenoScreen's MLST service offer is a complete package of services realized from extracted DNA or thermolysate. This includes the PCRs realization, the sequencing on a capillary sequencer, the sequences analysis and the strain assignment..

Multiple Loci VNTR Analysis (MLVA)

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The MLVA (Multiple Loci VNTR Analysis) is a genetic analysis method of VNTR (variable number of tandem repeats) pattern polymorphism based on their count. During a first step, each targeted VNTR locus is amplified by PCR with flanking-region specific primers. The fragments obtained are then separated according to their size by electrophoresis on capillary sequencer. The genotyping of each locus and the results compiled from each sample allow to assign to the strain a specific numerical code for a subspecies.

The MLVA approach allows to characterize a microorganism whose species is known, to identify, at least, the subspecies through allele typing and comparison to the MLVA databases. This method is more selective than the MLST method and does not require a DNA sequencing step. It thus makes it possible to differentiate close subspecies (clonal species).

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Le MLVA par GenoScreen

GenoScreen's MLVA service offer is a complete package of services realized from extracted DNA or thermolysate. This includes the PCRs realization, the electrophoresis on a capillary sequencer, the data analysis and the strain assignment. 

The analysis can be carried out on the basis of a personalised typing scheme tailored to your needs, in line with the data available for your species. 

This technique is based on the variation of a single nucleotide at a given position (Single Nucleotide Polymorphisms), which can lead to functional changes in the genes and proteins resulting from the modified amino acid sequence. By offering genome-wide analysis, GenoScreen can help you identify unknown genotype-phenotype associations or genetic variations that are already known.

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In what context should microsatellite genotyping be carried out?

Since SNPs can be found in any type of genome, genotyping them can be of interest in a wide range of applications. Our team is used to working on a wide range of problems, and can help you choose the best technique to meet your needs.

GenoScreen offer

Our services are tailored to your needs. We can use assays already available in databases, or identify and target SNPs of interest from sequencing data (with or without a reference genome).

Genotyping by qPCR

qPCR is used to identify genetic variations such as SNPs by specifically amplifying DNA sequences. This genotyping is based on methods such as TaqMan probes, which offer precise discrimination of alleles, or KASPar competitive PCR, which is more economical for large-scale or recurrent analyses.

These two methods for identifying genetic variants operated by GenoScreen will be proposed to you depending on the purpose and recurrence of your project. Our team can also help you design a genotyping kit.

 

 

A tailored approach to meet your needs

We work with you to plan all the stages required to bring your project to a successful conclusion, whatever your constraints in terms of number of samples or targets, timeframe and cost.

 

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Genotyping by sequencing

 

These technologies are commonly used in population genetics, conservation biology and ecology projects. It is not necessary to have access to a reference genome to carry out the analysis.

The data obtained can be used to identify SNPs as genetic markers, perform population structure and/or phylogenetic analyses.

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Sanger

Thanks to its reliability and accuracy, Sanger sequencing remains the reference method when it comes to verifying uncertain qPCR results or confirming mutations observed by high-throughput sequencing.

This technique is based on the amplification of short DNA sequences composed of repeating units of a few nucleotides. Because of their polymorphism, microsatellite markers can be used to study genetic diversity within a species or population. GenoScreen has developed almost 20 years of expertise in the study and analysis of microsatellites, enabling the development of markers for more than 500 species.

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When should microsatellite genotyping be used?

GenoScreen offer

 

 

 

Use of existing data

As microsatellite markers are molecular reference tools, there are many publications and previous works available today that we can draw on to meet your needs. We can also use existing data on closely related species (cross-amplification) or from high-throughput sequencing.

Whenever markers are already available for your species of interest, we can use the amplification conditions developed or develop new ones to, for example, group together markers from different publications or use only some of them. There are several possible scenarios:

Features

Multiplex development

(several pairs of primers)

From previously published publications / markers

Cross-amplification

Custom

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Development of de novo markers

 GenoSat® - GenoScreen offer

Geno Sat^® is a standardised, simple and modular solution for all types of eukaryotic species. Since 2009, our solution has already been used on more than 500 different species as part of phylogenetic studies, biodiversity analyses and conservation programmes. This solution enables :

• Production of microsatellite libraries using high-throughput sequencing,
• Development of polymorphic markers of interest
• Population genotyping

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Geno Sat®

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Using 1 µg of genomic DNA and Illumina MiSeq high-throughput sequencing technology, GenoSat® can produce bioinformatically validated primer pairs within 2 months, framing the microsatellites identified for your species.

Geno Sat® Pack

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This modular solution lets you both biologically validate a batch of primer pairs (after in silico validation), and study the polymorphism of successfully amplified pairs.

Geno Sat® Pack +

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The Geno Sat® Pack + solution is our most comprehensive offering for optimising multiplexes and/or genotyping large populations using polymorphic microsatellite markers of interest.

Discover the list of species analysed with Geno Sat® below:

 

Fragment analysis

Equipped with ABI 3730XL capillary sequencers, we can respond rapidly to your needs for fragment analysis (microsatellites, SSR, STR, etc.) using your labelled PCR products supplied in 96-well plates.

We offer sliding scale prices depending on the number of plates received per batch and migrated with the same size marker.

It is also possible to send us plates ready to be migrated using the same principle, already containing the Hi-Di Formamide + size marker mix with your sample.

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The identification and genotyping are essential in the discovery of genetics variations, specially in the studies of the populations and pathologies.

These analyses are extremely informative for many key scientific activities:

• Food traceability.
• Biomarker development and validation (screening/diagnostics, functional genomics, trait prediction models, etc.).
• Studies of genetic diversity and evolution (phylogeny, etc.).
• Identification of individuals (criminals, bodies) or links between individuals (paternity tests).
• Genomic selection (agriculture, animal husbandry, etc.).

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