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Genetics / Genomics JoVE (Open Access) Citable · DOI

Infinium Assay for Large-scale SNP Genotyping Applications

DOI: 10.3791/50683-v
What you'll learn
  • ✓ Perform large-scale SNP genotyping using Illumina Infinium assay workflow
  • ✓ Execute DNA amplification, fragmentation, and hybridization procedures
  • ✓ Interpret genotype data for disease and phenotype association studies
Protocol

A protocol is described that uses Illumina's Infinium assays to perform large-scale genotyping. These assays can reliably genotype millions of SNPs across hundreds of individual DNA samples in three days. Once generated, these genotypes can be used to check for associations with a variety of different diseases or phenotypes.

Difficulty
advanced
Total time
~3 days for hundreds of samples
Biosafety
BSL-1

Steps

1
Amplify genomic DNA using whole genome amplification

Perform isothermal DNA amplification to generate sufficient template from genomic DNA samples for subsequent genotyping analysis.

▶ 01:03
2
Fragment and precipitate amplified DNA product

Enzymatically fragment the amplified DNA and precipitate the product for purification before hybridization steps.

▶ 02:51
3
Hybridize SNP probes to fragmented DNA targets

Incubate fragmented DNA with SNP-specific oligonucleotide probes on Infinium BeadChip arrays under controlled hybridization conditions.

▶ 03:47
4
Prepare fluorescent staining reagents and solutions

Mix and prepare staining dyes and extension nucleotides for visualization and polymerase extension of hybridized SNP probes.

▶ 06:42
5
Perform single-base extension and fluorescent staining

Extend hybridized probes with fluorescently labeled nucleotides and apply staining reagents to generate genotype-specific signals.

▶ 09:29
6
Wash excess reagents and seal BeadChip array

Remove unincorporated stain and reagents through washing steps, then seal the array for scanning and data acquisition.

▶ 10:29
7
Scan array and analyze SNP genotype data

Capture fluorescent images from millions of SNPs across all samples and process raw intensity data into genotype calls for association studies.

▶ 11:11
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