{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,25]],"date-time":"2026-09-25T20:59:29Z","timestamp":1790369969545,"version":"4.1.0"},"reference-count":41,"publisher":"Wiley","issue":"22","license":[{"start":{"date-parts":[[2023,9,28]],"date-time":"2023-09-28T00:00:00Z","timestamp":1695859200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32200496"],"award-info":[{"award-number":["32200496"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32071218"],"award-info":[{"award-number":["32071218"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["chemistry-europe.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["ChemBioChem"],"published-print":{"date-parts":[[2023,11,16]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>RNA\u2010guided protease activity was recently discovered in the type III\u2010E CRISPR\u2010Cas systems (Craspase), providing a novel platform for engineering a protein probe instead of the commonly used nucleic acid probe in nucleic acid detection assays. Here, by adapting a fluorescence readout technique using the affinity\u2010 and fluorescent protein dual\u2010tagged Csx30 protein substrate, we have established an assay monitoring Csx30 cleavage by target ssRNA\u2010activated Craspase. Four Craspase\u2010based nucleic acid detection systems for genes from severe acute respiratory syndrome coronavirus 2 (SARS\u2010CoV\u20102), norovirus, and the influenza virus (IFV) were reconstituted with demonstrated specificity. The assay could reliably detect target ssRNAs at concentrations down to 25\u2005pM, which could be further improved approximately 15\u2009000\u2010fold (ca. 2\u2005fM) by incorporating a recombinase polymerase isothermal preamplification step. Importantly, the species\u2010specific substrate cleavage specificity of Craspase enabled multiplexed diagnosis, as demonstrated by the reconstituted composite systems for simultaneous detection of two genes from the same virus (SARS\u2010CoV\u20102, <jats:italic>spike<\/jats:italic> and <jats:italic>nsp12<\/jats:italic>) or two types of viruses (SARS\u2010CoV\u20102 and IFV). The assay could be further expanded by diversifying the fluorescent tags in the substrate and including Craspase systems from various species, thus potentially providing an easily adaptable platform for clinical diagnosis.<\/jats:p>","DOI":"10.1002\/cbic.202300401","type":"journal-article","created":{"date-parts":[[2023,9,14]],"date-time":"2023-09-14T23:02:37Z","timestamp":1694732557000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":6,"title":["Nucleic Acid Detection through RNA\u2010Guided Protease Activity in Type III\u2010E CRISPR\u2010Cas Systems"],"prefix":"10.1002","volume":"24","author":[{"given":"Qiuqiu","family":"He","sequence":"first","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xinlong","family":"Lei","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuanjun","family":"Liu","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Dermatovenereology Tianjin Medical University General Hospital  154 Anshan Road Tianjin 300052 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Xiaoshen","family":"Wang","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nan","family":"Ji","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hang","family":"Yin","sequence":"additional","affiliation":[{"name":"Department of Pharmacology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Huiping","family":"Wang","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Dermatovenereology Tianjin Medical University General Hospital  154 Anshan Road Tianjin 300052 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heng","family":"Zhang","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1322-8288","authenticated-orcid":false,"given":"Guimei","family":"Yu","sequence":"additional","affiliation":[{"name":"The Province and Ministry Co-sponsored Collaborative Innovation Center for Medical Epigenetics Key Laboratory of Immune Microenvironment and Disease (Ministry of Education) Tianjin Medical University General Hospital Tianjin Medical University  Tianjin 300070 P. R. China"},{"name":"Department of Biochemistry and Molecular Biology School of Basic Medical Sciences Tianjin Medical University  Tianjin 300070 P. R. China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2023,9,28]]},"reference":[{"key":"e_1_2_9_1_1","doi-asserted-by":"publisher","DOI":"10.1136\/bmjgh-2018-001179"},{"key":"e_1_2_9_2_1","unstructured":"\u00a0"},{"key":"e_1_2_9_3_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2017.11.032"},{"key":"e_1_2_9_4_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro.2016.184"},{"key":"e_1_2_9_5_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41579-019-0299-x"},{"key":"e_1_2_9_6_1","unstructured":"\u00a0"},{"key":"e_1_2_9_7_2","doi-asserted-by":"publisher","DOI":"10.1038\/nature19802"},{"key":"e_1_2_9_8_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.aar6245"},{"key":"e_1_2_9_9_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2018.10.052"},{"key":"e_1_2_9_10_1","unstructured":"\u00a0"},{"key":"e_1_2_9_11_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.aam9321"},{"key":"e_1_2_9_12_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.aaq0179"},{"key":"e_1_2_9_13_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-021-25977-7"},{"key":"e_1_2_9_14_1","unstructured":"\u00a0"},{"key":"e_1_2_9_15_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.abk2718"},{"key":"e_1_2_9_16_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-021-03886-5"},{"key":"e_1_2_9_17_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2022.05.003"},{"key":"e_1_2_9_18_1","unstructured":"\u00a0"},{"key":"e_1_2_9_19_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.add5064"},{"key":"e_1_2_9_20_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41564-022-01256-z"},{"key":"e_1_2_9_21_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.add7450"},{"key":"e_1_2_9_22_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41594-022-00894-5"},{"key":"e_1_2_9_23_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.add7347"},{"key":"e_1_2_9_24_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2022.10.007"},{"key":"e_1_2_9_25_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-022-35275-5"},{"key":"e_1_2_9_26_2","doi-asserted-by":"publisher","DOI":"10.1038\/s41422-022-00738-3"},{"key":"e_1_2_9_27_2","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.81678"},{"key":"e_1_2_9_28_1","unstructured":"\u00a0"},{"key":"e_1_2_9_29_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2020.10.004"},{"key":"e_1_2_9_30_2","doi-asserted-by":"publisher","DOI":"10.1002\/jmv.26222"},{"key":"e_1_2_9_31_1","unstructured":"\u00a0"},{"key":"e_1_2_9_32_2","doi-asserted-by":"publisher","DOI":"10.1053\/j.spid.2004.07.004"},{"key":"e_1_2_9_33_2","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2020.00961"},{"key":"e_1_2_9_34_1","volume":"8","author":"Dziabowska K.","year":"2018","journal-title":"Biosensors (Basel)"},{"key":"e_1_2_9_35_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pbio.0040204"},{"key":"e_1_2_9_36_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkac1151"},{"key":"e_1_2_9_37_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41551-020-00603-x"},{"key":"e_1_2_9_38_1","unstructured":"\u00a0"},{"key":"e_1_2_9_39_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2021.01.044"},{"key":"e_1_2_9_40_2","doi-asserted-by":"publisher","DOI":"10.3389\/fimmu.2021.711997"},{"key":"e_1_2_9_41_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41589-022-01135-y"}],"container-title":["ChemBioChem"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/chemistry-europe.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/cbic.202300401","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T10:59:03Z","timestamp":1760266743000},"score":1,"resource":{"primary":{"URL":"https:\/\/chemistry-europe.onlinelibrary.wiley.com\/doi\/10.1002\/cbic.202300401"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,9,28]]},"references-count":41,"journal-issue":{"issue":"22","published-print":{"date-parts":[[2023,11,16]]}},"alternative-id":["10.1002\/cbic.202300401"],"URL":"https:\/\/doi.org\/10.1002\/cbic.202300401","archive":["Portico"],"relation":{},"ISSN":["1439-4227","1439-7633"],"issn-type":[{"value":"1439-4227","type":"print"},{"value":"1439-7633","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,9,28]]},"assertion":[{"value":"2023-05-29","order":0,"name":"received","label":"Received","group":{"name":"publication_history","label":"Publication History"}},{"value":"2023-09-28","order":3,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}],"article-number":"e202300401"}}