TaqMan is a commonly used SNP genotyping technique invented by Life Technologies. Each TagMan Genotyping Assay contains two primers for amplifying the sequence of interest and two allele-specific probes for allele detection.
The probes have a fluorophore linked to their 5’ end and a quencher molecule linked to their 3’ end. While the probe is intact, the proximity of the quencher dye to the reporter dye suppresses the reporter fluorescence. During the PCR amplification step, if the allele-specific probe is perfectly complementary to the SNP allele, it will hybridise to the target DNA segment and then get degraded by the 5’-nuclease activity of the Taq polymerase. The degradation of the probe results in the separation of the fluorophore from the quencher molecule, generating a detectable fluorescent signal. If the allele-specific probe is not perfectly complementary, it will have a lower melting temperature and will not hybridise to target as efficiently. This prevents the nuclease from acting on the probe.
The TaqMan platform is most appropriate for projects of 10s of SNPs and 1000s of Samples.
Technique Highlights
– Most appropriate for projects of medium size (100-1000 samples) being genotyped for a small number of SNPs (1-10)
– Simple and robust chemistry
– A large number of pre-validated assays are commercially available
Sample Requirement
Sample Type: Genomic DNA
Amount: DNA >= 2 ug; concentration >= 25 ng/ul
Purity: OD260/280 = 1.7~2.0
Sample without severe degradation
Service Workflow
1. Cusomtized sequencing experimental design
2. Sample receiving
3. DNA extraction and QC
4. Taqman assay
5. Project closure report
Deliverables
1. Experimental conditions including information of primer and probe sequences, reagent names, device models, protocols.
2. Clustering graphs and genotyping tables of each SNP locus in individual sample.
