What is Genotyping?

Single nucleotide polymorphisms (SNPs) are a widely studied form of genetic variation in humans, SNPs are single base-pair changes in DNA that occur at specific locations in the genome. For example, most individuals might carry the C nucleotide at a specific base position, but in some individuals, this is replaced by an A. This variation is known as a SNP.

SNPs can explain traits such as eye color and inherited diseases like cystic fibrosis and sickle cell anemia. They also act as markers indicating a risk of developing complex diseases like diabetes and Alzheimer's disease. SNP genotyping can accelerate personalized medicine by predicting an individual's risk of developing certain diseases and designing targeted therapies specific to the genetic basis of the disease.

Insertion or deletion polymorphisms (indels) are the insertion or deletion of a small number of bases in a genome and are the second most common class of mutation in the human genome. Indels can significantly affect gene expression, causing frameshift mutations unless the indel length is a multiple of three. These mutations have been implicated in various genetic diseases.

Copy number variants (CNVs) are variations in the number of copies of a specific gene between individuals. They affect a significant portion of the genome (approximately 12% of the human genome) and include deletions, duplications, and other complex genotyping patterns. CNVs can influence gene expression and are associated with specific phenotypes and diseases, such as microdeletion and microduplication syndromes.

There are many methods for detecting novel and known SNPs, including DNA sequencing, mass spectrometry, molecular beacons, SNP microarrays, and PCR-based methods. SNP detection can be categorized into two sub-groups: SNP discovery and SNP screening. SNP discovery involves identifying new SNPs in targeted areas or on a genome-wide scale, while SNP screening pertains to known SNPs and involves genotyping individuals or determining if a particular SNP is involved in producing a certain characteristic.

Disease association

Genome-wide association studies (GWAS) can identify connections between SNPs and common disease risk by comparing the polymorphisms across different populations (one healthy and one diseased). GWAS can also help unravel the biological processes underlying disease states by identifying potential causal factors [1–4].

Population genomics

SNPs have implications for evolutionary biology and can help identify genetic variations that underlie phenotypic differences between healthy individuals. Understanding normal genetic variation across different populations helps us understand how different groups have evolved and diverged, with implications for protecting species against future environmental challenges.

Trait selection in agriculture

Genetic variation is particularly beneficial in agriculture, where trait selection in plants and livestock has been used for centuries to increase yield and quality. Modern selective breeding relies heavily on molecular biology techniques, including SNP genotyping, to detect functionally relevant genetic changes and design intelligent breeding programs.

Microorganisms

SNP genotyping can discriminate between bacterial isolates and characterize strains of antibiotic resistance. This is relevant in both clinical and agricultural research and has been used to study a range of infectious diseases in humans and plants [5,6].

Real-time PCR is an easy, accurate, and scalable method for screening known SNPs. TaqMan SNP genotyping assays use 5'-nuclease chemistry to determine whether a given SNP is present in a sample. These assays include a primer pair to amplify the target area and two allele-specific probes to detect target SNP alleles and report the genotypes of samples.

Millions of predesigned TaqMan SNP genotyping assays are available for human and mouse subjects. Custom TaqMan SNP genotyping assays can also be designed for other species using the online Custom Assay Design Tool. These assays can be applied to both animal and plant species, including polyploid plants, providing unparalleled versatility.

TaqMan assays demonstrate speed, specificity, sensitivity, ease of use, and reproducibility, detecting sequences using tiny DNA concentrations in the nanogram range. Utilizing TaqMan 5'-nuclease chemistry, these assays include a primer pair to amplify the target area and two allele-specific probes to detect target SNP alleles and report sample genotypes. TaqMan SNP genotyping assays have been used to identify variants associated with blood disorders [7] and many other diseases, including evaluating almost 1,200 human DNA samples for high-throughput ApoE genotyping to assess Alzheimer's disease risk [8]. These assays can genotype transgenic mice [9], helping to create pure lines and reduce experimental variability. They also have been applied to evaluate the origin of human and animal parasites, valuable for medicine and agriculture [10]. With millions of predesigned assays available for human and mouse subjects, and the ability to design custom assays for other species using the online Custom Assay Design Tool, TaqMan SNP genotyping assays can be applied to both animal and plant species, including polyploid plants.

Each assay aligns uniquely with the genome to specifically detect either allele in the sequence of interest and can be used on a range of real-time PCR instrument types, providing unparalleled versatility. TaqMan assays are an ideal and cost-effective method for investigating SNPs in large populations.

Explore:TaqMan SNP genotyping assays

1. Visscher PM, Wray NR, Zhang Q, et al. (2017) 10 Years of GWAS Discovery: Biology, Function, and Translation. Am J Hum Genet Jul 6;101(1):5–22.
2. Gaj P, Maryan N, Hennig EE, et al. (2012) Pooled sample-based GWAS: A cost-effective alternative for identifying colorectal and prostate cancer risk variants in the Polish population. PLoS ONE 7(4):e35307.
3. Figueroa JD, Ye Y, Siddiq A, et al. (2014) Genome-wide association study identifies multiple loci associated with bladder cancer risk. Hum Mol Gen 23(5):1387–1398.
4. Reddy MPL, Wang H, Liu S, et al. (2011) Association between type 1 diabetes and GWAS SNPs in the southeast US Caucasian population. Genes Immun 12(3):208–212.
5. Sengstake S, Bablishvili N, Schuitema A et al. (2014) Optimizing multiplex SNP-based data analysis for genotyping of Mycobacterium tuberculosis isolates, BMC Genomics 15(1):572.
6. Rathnayake I, Hargreaves M, Huygens F. (2011) SNP diversity of Enterococcus faecalis and Enterococcus faecium in a South East Queensland waterway, Australia, and associated antibiotic resistance gene profiles. BMC Microbiol 11(1):201.
7. Teh L-K, Lee T-Y, Tan JAMA, et al. (2015) The use of Taqman genotyping assays for rapid confirmation of β-thalassaemia mutations in the Malays: Accurate diagnosis with low DNA concentrations. Int J Lab Hematol 37(1):79–89. doi: 10.1111/ijlh.12240.
8. Zhong L, Xie Y-Z, Cao T-T et al. (2016) A rapid and cost-effective method for genotyping apolipoprotein E gene polymorphism. Mol Neurodegener 11(1):2.
9. Vaisman BL (2013) Genotyping of transgenic animals by real-time quantitative PCR with TaqMan probes. Methods Mol Biol 1027:233–251.
10. Burnet JB, Ogorzaly L, Tissier A, et al. (2013) Novel quantitative TaqMan real-time PCR assays for detection of Cryptosporidium at the genus level and genotyping of major human and cattle-infecting species. J Appl Microbiol 114(4):1211–1222.