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. 2021 Jul;5(7):713-725.
doi: 10.1038/s41551-021-00706-z. Epub 2021 Apr 5.

Discrimination of single-point mutations in unamplified genomic DNA via Cas9 immobilized on a graphene field-effect transistor

Sarah Balderston  1   2 , Jeffrey J Taulbee  3 , Elizabeth Celaya  2 , Kandace Fung  1 , Amanda Jiao  2 , Kasey Smith  1 , Reza Hajian  1   2 , Giedrius Gasiunas  4   5 , Simonas Kutanovas  4 , Daehwan Kim  6 , Jonathan Parkinson  2 , Kenneth Dickerson  2 , Juan-José Ripoll  2 , Regis Peytavi  2 , Hsiang-Wei Lu  1   2 , Francie Barron  2 , Brett R Goldsmith  2 , Philip G Collins  3 , Irina M Conboy  6 , Virginijus Siksnys  4   5 , Kiana Aran  7   8   9
Affiliations

Discrimination of single-point mutations in unamplified genomic DNA via Cas9 immobilized on a graphene field-effect transistor

Sarah Balderston et al. Nat Biomed Eng. 2021 Jul.

Abstract

Simple and fast methods for the detection of target genes with single-nucleotide specificity could open up genetic research and diagnostics beyond laboratory settings. We recently reported a biosensor for the electronic detection of unamplified target genes using liquid-gated graphene field-effect transistors employing an RNA-guided catalytically deactivated CRISPR-associated protein 9 (Cas9) anchored to a graphene monolayer. Here, using unamplified genomic samples from patients and by measuring multiple types of electrical response, we show that the biosensors can discriminate within one hour between wild-type and homozygous mutant alleles differing by a single nucleotide. We also show that biosensors using a guide RNA-Cas9 orthologue complex targeting genes within the protospacer-adjacent motif discriminated between homozygous and heterozygous DNA samples from patients with sickle cell disease, and that the biosensors can also be used to rapidly screen for guide RNA-Cas9 complexes that maximize gene-targeting efficiency.

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