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Custom SNP Genotyping

Single nucleotide polymorphisms (SNPs) represent the most frequent type of DNA variation. Utilising a variety of platforms, AGRF offers a custom SNP service which can be tailored to meet specific project requirements. AGRF's custom SNP Genotyping services are based on the following:

Sequenom platform which has the capacity to analyse SNPs ranging from only one SNP to >500 SNPs to genotype for virtually any number of samples.
Illumina Golden Gate Genotyping System which offers custom SNP panels from 96 - to 1536-plex and has the capacity to analyse SNPs ranging from 300 to 10 000 SNPs.

Custom SNP: Sample Submission Requirements

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Illumina Custom GoldenGate Genotyping

AGRF utilises the high through-put illumina iScan System to deliver the GoldenGate Genotyping Assay, a flexible and cost effective SNP assay. This assay is used to interrogate either 96 loci or between 384-1536 SNPs (in multiples of 96), simultaneously making it ideal for large scale association studies. GoldenGate panels can be processed on 12-, and 32-sample Universal BeadChips.

Using the GoldenGate Assay technology, the AGRF provides the following services:

Custom GoldenGate Panels: These panels can be custom designed to meet the researchers’ needs, providing greater flexibility with user defined content across a wide range of genomes. For GoldenGate Custom Panels, AGRF can assist and facilitate in the selection of markers based on your region and density requirements. The final conversion success rate is influenced by:

Stringent QC criteria adhered to by illumina during manufacture of GoldenGate custom panels
The sequence nature of the chosen loci
Analysis criteria

GoldenGate Focused Content Panels encompass a variety of BeadChips that have been optimised for screening specific interest areas. Also available using the GoldenGate assay are standard panels for mouse and human linkage:

Please note that AGRF recommend selecting validated loci and/or loci with high design scores.

Illumina Custom GoldenGate Genotyping Process

Infinium iSelect HD Custom Genotyping Panels

The Infinium iSelect HD Genotyping BeadChip is extremely flexible, allowing researchers to select practically any SNP in the genome for any species of interest. With iSelects high throughput capabilities, it offers the ability to simultaneously process up to 24-samples and screen between 3,000 and 200,000 custom SNPs. The iSelect BeadChips utilise Infinium chemistry for performing large scale custom genotyping, such as fine mapping.

The Infinium iSelect HD Genotyping BeadChips can be used to additionally include up to 60,800 custom SNPs per sample for both the semi-custom HumanHap1M-Duo+ and the HumanHap550+ BeadChips or up to 200,000 markers on the HumanOmniExpress+.

Please note that the minimum order for iSelect BeadChips is for 1,152 samples.

Infinium iSelect HD Custom Genotyping Panels Process

Sequenom Custom SNP Genotyping

AGRF's Sequenom custom SNP genotyping service is run on the Sequenom MassARRAY system and utilises a homogenous MassExtend (hME – single base extension) reaction termed iPLEX GOLD, for analysis of up to 36 SNP's in a single multiplex. SNP assays are designed by the AGRF based on provided SNP sequences (RS Numbers). Oligonucleotides are then obtained by the AGRF and used to process samples in multiplex format. Following the MassExtend reaction, oligonucleotides are printed onto specialised Spectro CHIPs and anaylsed by the Sequenom mass spectrometer. This platform has the ability to replex SNP panels so SNPs can be dropped or added at a later date.

Sequenom Homogenous MassExtend Bulletin (PDF)

Sequenom iPlex Application Note (PDF)

Sequenom Custom SNP Genotyping Service Process

Sequenom Custom SNP Allelotyping

AGRF offers a rapid and accurate custom SNP validation using a sample-pooling technique called allelotyping. The allelotyping technique rapidly screens and confirms the presence of a SNP and its allelic frequency in cohort populations. The accuracy of the Sequenom MassARRAY technology is able to determine SNP allele frequencies with high precision in pooled samples, thereby replacing hundreds of individual measurements with one consolidated analysis. The MassARRAY has the capacity to pre-evaluate SNPs, which means only SNPs that prove informative in the pool-to-pool comparison are included in individual genotyping projects, significantly reducing the total workload for individual genotyping. Allelotyping may also be used when looking for allele bias in cDNA.

AGRF allelotyping service requires a SNP sequence with a minimum 50 bases either side of the SNP site and extracted DNA pools (screening 300-500 individuals per patient pool is recommend for greatest statistical relevance). SNP assays are then designed by the AGRF and PCR oligonucleotides are obtained. PCR utilises the homogenous MassExtend (hME – single base extension) reaction termed iPLEX GOLD, with pooled samples then being processing on a mass spectrometer. The mass spectrometer provides a high level of sensitivity, capable of detecting allele frequencies as low as 3% on average.