Most methods for the detection of nucleic acids require many reagents and expensive and bulky instrumentation. Here, we report the development and testing of a graphene-based field-effect transistor that uses clustered regularly interspaced short palindromic repeats (CRISPR) technology to enable the digital detection of a target sequence within intact genomic material. Termed CRISPR–Chip, the biosensor uses the gene-targeting capacity of catalytically deactivated CRISPR-associated protein 9 (Cas9) complexed with a specific single-guide RNA and immobilized on the transistor to yield a label-free nucleic-acid-testing device whose output signal can be measured with a simple handheld reader. We used CRISPR–Chip to analyse DNA samples collected from HEK293T cell lines expressing blue fluorescent protein, and clinical samples of DNA with two distinct mutations at exons commonly deleted in individuals with Duchenne muscular dystrophy. In the presence of genomic DNA containing the target gene, CRISPR–Chip generates, within 15 min, with a sensitivity of 1.7 fM and without the need for amplification, a significant enhancement in output signal relative to samples lacking the target sequence. CRISPR–Chip expands the applications of CRISPR–Cas9 technology to the on-chip electrical detection of nucleic acids.
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The authors declare that all data supporting the findings in this study are available within the paper and its Supplementary Information files.
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We acknowledge Cardea Bio for our use of their Agile R100 reader technology. We thank J. Corn (University of California, Berkeley) for providing us with the HEK-BFP cells. This work was primarily supported by Keck Start-up funding to the Aran Lab, by an Open Philanthropy Research Gift and by the Rogers Family Foundation to I.C.
K.A. is a co-founder of Nanosens Innovations, and R.P. is Vice President of Technology Development in the same company. The other authors declare no competing interests.
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Hajian, R., Balderston, S., Tran, T. et al. Detection of unamplified target genes via CRISPR–Cas9 immobilized on a graphene field-effect transistor. Nat Biomed Eng 3, 427–437 (2019). https://doi.org/10.1038/s41551-019-0371-x
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