Low-cost allele-specific PCR for saliva-based SNP detection

Overview

This innovative solution offers a low-cost allele-specific PCR method for detecting single nucleotide polymorphisms (SNPs) using saliva samples. The method utilizes heat denaturation for DNA extraction followed by allele-specific PCR, making it a cost-effective alternative to sequencing-based assays. Inspired by the covidSHIELD assay, this approach is adapted for SNP detection with high throughput and accuracy, suitable for large-scale genetic testing.

Technical specifications

• Saliva Sampling and DNA Extraction: Saliva samples are heat-denatured at 95°C for 30 minutes to extract DNA.
• PCR Amplification: The denatured sample is mixed with TBE and Tween-20, then amplified using allele-specific primers designed to inhibit amplification in SNP-negative samples.
• Detection Methods: Results can be analyzed via qPCR, dPCR, or conventional PCR with gel electrophoresis, allowing detection of multiple SNPs through fluorescent probes or differential amplicon lengths.
• Multiplexing Capability: Capable of detecting 2-3 SNPs per assay with qPCR and 5 or more with dPCR or gel electrophoresis.

Technology readiness level

This technology is at TRL 6, having been demonstrated in a relevant environment with plans for further validation on DNA stability and sensitivity using various storage conditions.