Understanding genotyping


Genotyping is the process of determining the DNA sequence—the genotype—at specific positions within a gene of an individual. Sequence variations can be used as markers in linkage and association studies to determine genes relevant to specific traits or disease.

pipette tips used in genotyping analysis

Identifying sequence variation

Genotyping is used to identify variation at specific positions in the DNA sequence of any biological species, from microorganisms to humans. This variation, the genotype, occurs naturally and can be considered a genetic fingerprint of an individual. It is identified as distinct when compared to a reference sequence derived from the general population or a defined subgroup, and can differ from the reference sequence in numerous ways. Types of variation include single base changes (commonly referred to as single nucleotide variants and polymorphisms, or SNVs and SNPs), insertions and deletions (indels), and even the number of gene copies (copy number variation, or CNV).

SNPs are the most common type of sequence variant; they are typically defined as SNVs that occur at >1% in the population. Based on the number of SNPs cataloged in Build 149 of the SNP database, dbSNP, maintained by the National Center for Biotechnology Information (NCBI) [1] and a genome size of 3.4 x 109 bp [2], the human genome should contain a SNP approximately once every 22 bases! Other common model systems show a similarly high frequency of SNPs [3].

A series of adjacent SNPs from an individual represent a haplotype and can serve as a signature for a specific phenotypic trait, such as fruit production or certain cancers (Figure 1).

Figure 1. SNPs contributing to haplotypes in individual sequences. An example of SNPs (A or G, G or T, and A or T), in this case at the 3 variant positions in this stretch of DNA sequence. The SNPs of each individual sequence are combined to denote 3 distinct haplotypes (AGT, GTA, AGA).

Associating sequence variation to specific traits

By comparing sequence variations among individual plants, animals, or humans, researchers can identify heritable genes relevant to specific traits. These unique differences can be used as markers in linkage and association studies to:

  • Map gene function and genetic variation
  • Identify gene variants associated with a unique phenotype or disease; where SNPs serve as genetic markers (biomarkers)
  • Match effective medical treatments to populations with specific genotypes (personalized medicine)
  • Facilitate animal breeding, such as the selection of a desired genotype from a cross hybrid
  • Trace ancestry or the origins of disease; map evolution, such as phylogenetic relationships between species; or test for family relationship for inheritance or paternity
  • Carry out population association studies, such as genome-wide linkage analysis, which use SNPs as markers
  • Perform forensics, using genotyping to identify a specific individual
  • Do pathogen typing and resistance screening
  • Monitor biodiversity

As mentioned above, there are also other types of information collected apart from sequence differences, such as copy number variation (CNV).

Technologies used to study genotyping

There are different approaches to SNP genotyping with the number of samples, the number of genotypes to be tested, and the amount of sample material available, all factoring into choice of technology. Methods include whole genome analysis by NGS or microarrays, or more targeted analysis using qPCR, dPCR, or targeted sequencing.

Experimental design can be low throughput vs. highly multiplexed (look at numerous SNPs simultaneously). Common analysis methods include end-point or quantitative PCR (qPCR), targeted sequencing, bead or microarray analysis, and even mass spectrometry.

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Overview of genotyping terms

Learn more about the language used in genotyping studies.

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IDT genotyping solutions

Genotyping by PCR

Polymerase chain reaction (PCR) is a commonly used genotyping technique. Often employing a primer-pair and target-specific fluorescent probe, quantitative PCR (qPCR) can be a sensitive and specific way to detect SNPs. IDT offers a complete SNP-typing solution with predesigned assays, as well as complementary easy- and ready-to-use reagent mixes. For other applications, modified probes are available that can be incorporated into custom qPCR assays.

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Genotyping by targeted sequencing

Targeted sequencing uses deep sequencing to detect known and novel variants within your region of interest. Thus, it can be used as a method for SNP and mutation detection, and gene structural analysis.

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Get started with genotyping

If you choose to use qPCR technology, the rhAmp SNP Genotyping System provides a fully integrated, high performing solution for out-of-the-box convenience. For user-defined methods, order custom probes with modifications, such as Affinity Plus qPCR locked nucleic acid Probes, that increase probe stability and enable designs within difficult sequences and with selective target identification.

rhAmp SNP Genotyping System

The rhAmp SNP Genotyping System is a fully integrated genotyping solution that includes an extensive predesigned assay collection, a custom design tool, optimized reagent mixes, and optional synthetic control templates. SNP detection may be performed on any commonly available qPCR instrument.

Precise and easy-to-use, the rhAmp SNP Genotyping System offers an out-of-the-box solution for SNP genotyping studies for small discovery or large screening projects.

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Affinity Plus qPCR Probes

Use Affinity Plus qPCR Probes for SNP genotyping, transcript variant identification, and more sensitive target detection in challenging samples (FFPE tissue, biofluids). The Affinity Plus bases used in these qPCR probes are locked nucleic acid monomers. When incorporated into a probe, they impart heightened structural stability, leading to increased hybridization melt temperature (Tm).

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PACE™ SNP Genotyping Assays

PACE™ SNP genotyping uses competitive, allele-specific PCR and a simple, easily detected fluorescent readout for bi-allelic genotyping. Obtain primer sets from IDT for cost-effective, high-throughput assays.

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rhAmpSeq amplicon sequencing system

The rhAmpSeq system enables highly accurate amplicon sequencing on Illumina next generation sequencing (NGS) platforms. Whether you are investigating thousands of targets or a few, the fast and easy rhAmpSeq workflow generates NGS-ready amplicon libraries for deep, targeted resequencing.

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  1. NCBI dbSNP Build 149 (Nov 7, 2016) www.ncbi.nlm.nih.gov/dbvar/content/org_summary/ [Accessed Dec 19, 2016].
  2. Gregory TR. (2005) Animal genome size database. www.genomesize.com/ [Accessed Dec 19, 2016].
  3. Prediger E. (2017) Consider SNPs when designing PCR and qPCR assays, [Online] Coralville, Integrated DNA Technologies. [Accessed Jan 6, 2020].