Array-based Dynamic Allele Specific Hybridization (Array-DASH): optimization-free microarray processing for multiple simultaneous genomic assays.

Gibson, Spencer J; Zahra, Nathalie; Freeman, Peter J; Howard, Caroline; Lancaster, Owen; Veal, Colin; Fontdevila, Maria Casadellà ; Paredes, Roger; Noguera-Julian, Marc; Slater, Adrian; Brookes, Anthony J

We report proof-of-principle experiments regarding a dynamic microarray protocol enabling accurate and semi-quantitative DNA analysis for re-sequencing, fingerprinting and genotyping. Single-stranded target molecules hybridise to surface-bound probes during initial gradual cooling with high-fidelity. Real-time tracking of target denaturation (via fluorescence) during a 'dynamic' gradual heating phase permits 'melt-curve' analysis. The probe most closely matching the target sequence is identified based on the highest melting temperature. We demonstrated a >99% re-sequencing accuracy and a potential detection rate of 1% for SNPs. Experiments employing Hypericum ribosomal ITS regions and HIV genomes illustrated a reliable detection level of 5% plus simultaneous re-sequencing and genotyping. Such performance suggests a range of potential real-world applications involving rapid sequence interrogation, for example, in the Covid-19 pandemic. Guidance is offered towards the development of a commercial platform and dedicated software required to bring this technique into mainstream science.