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

Autor: Balderston S; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA.; Cardea, San Diego, CA, USA., Taulbee JJ; University of California, Irvine, Irvine, CA, USA., Celaya E; Cardea, San Diego, CA, USA., Fung K; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA., Jiao A; Cardea, San Diego, CA, USA., Smith K; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA., Hajian R; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA.; Cardea, San Diego, CA, USA., Gasiunas G; CasZyme, Vilnius, Lithuania.; Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius, Lithuania., Kutanovas S; CasZyme, Vilnius, Lithuania., Kim D; University of California, Berkeley, Berkeley, CA, USA., Parkinson J; Cardea, San Diego, CA, USA., Dickerson K; Cardea, San Diego, CA, USA., Ripoll JJ; Cardea, San Diego, CA, USA., Peytavi R; Cardea, San Diego, CA, USA., Lu HW; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA.; Cardea, San Diego, CA, USA., Barron F; Cardea, San Diego, CA, USA., Goldsmith BR; Cardea, San Diego, CA, USA., Collins PG; University of California, Irvine, Irvine, CA, USA., Conboy IM; University of California, Berkeley, Berkeley, CA, USA., Siksnys V; CasZyme, Vilnius, Lithuania.; Institute of Biotechnology, Life Sciences Center, Vilnius University, Vilnius, Lithuania., Aran K; Keck Graduate Institute, The Claremont Colleges, Claremont, CA, USA. karan@kgi.edu.; Cardea, San Diego, CA, USA. karan@kgi.edu.; University of California, Berkeley, Berkeley, CA, USA. karan@kgi.edu.
Jazyk: angličtina
Zdroj: Nature biomedical engineering [Nat Biomed Eng] 2021 Jul; Vol. 5 (7), pp. 713-725. Date of Electronic Publication: 2021 Apr 05.
DOI: 10.1038/s41551-021-00706-z
Abstrakt: 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.
(© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)
Databáze: MEDLINE
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