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Here, we present a CRISPR-based diagnostic assay that is robust to viral genome mutations and temperature, produces results fast, can be applied directly on nasopharyngeal (NP) specimens without RNA purification, and incorporates a human internal control within the same reaction. Specifically, we show that the use of an engineered AsCas12a enzyme enables detection of wildtype and mutated SARS-CoV-2 and allows us to perform the detection step with loop-mediated\u00a0isothermal amplification (LAMP) at 60-65\u2009\u00b0C. We also find that the use of hybrid DNA-RNA guides increases the rate of reaction, enabling our test to be completed within 30\u2009minutes. Utilizing clinical samples from 72 patients with COVID-19 infection and 57 healthy individuals, we demonstrate that our test exhibits a specificity and positive predictive value of 100% with a sensitivity of 50 and 1000 copies per reaction (or 2 and 40 copies per microliter) for purified RNA samples and unpurified NP specimens respectively.<\/jats:p>","DOI":"10.1038\/s41467-021-21996-6","type":"journal-article","created":{"date-parts":[[2021,3,19]],"date-time":"2021-03-19T09:05:27Z","timestamp":1616144727000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":207,"title":["An engineered CRISPR-Cas12a variant and DNA-RNA hybrid guides enable robust and rapid COVID-19 testing"],"prefix":"10.1038","volume":"12","author":[{"given":"Kean Hean","family":"Ooi","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3121-2201","authenticated-orcid":false,"given":"Mengying Mandy","family":"Liu","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie Wen Douglas","family":"Tay","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Seok Yee","family":"Teo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Pornchai","family":"Kaewsapsak","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shengyang","family":"Jin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6065-0000","authenticated-orcid":false,"given":"Chun Kiat","family":"Lee","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jingwen","family":"Hou","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0813-9640","authenticated-orcid":false,"given":"Sebastian","family":"Maurer-Stroh","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Weisi","family":"Lin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benedict","family":"Yan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3513-428X","authenticated-orcid":false,"given":"Gabriel","family":"Yan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yong-Gui","family":"Gao","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3627-5586","authenticated-orcid":false,"given":"Meng How","family":"Tan","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2021,3,19]]},"reference":[{"key":"21996_CR1","doi-asserted-by":"publisher","first-page":"270","DOI":"10.1038\/s41586-020-2012-7","volume":"579","author":"P Zhou","year":"2020","unstructured":"Zhou, P. et al. 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