{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,18]],"date-time":"2026-05-18T12:45:48Z","timestamp":1779108348016,"version":"3.51.4"},"reference-count":34,"publisher":"Wiley","issue":"1","license":[{"start":{"date-parts":[[2017,3,1]],"date-time":"2017-03-01T00:00:00Z","timestamp":1488326400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2017,3,1]],"date-time":"2017-03-01T00:00:00Z","timestamp":1488326400000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/www.springer.com\/tdm"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["R01GM107597"],"award-info":[{"award-number":["R01GM107597"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Quant. Biol."],"published-print":{"date-parts":[[2017,3]]},"abstract":"<jats:sec><jats:title>Background<\/jats:title><jats:p>The CRISPR\u2010Cas system is a widespread prokaryotic defense system which targets and cleaves invasive nucleic acids, such as plasmids or viruses. So far, a great number of studies have focused on the components and mechanisms of this system, however, a direct visualization of CRISPR\u2010Cas degrading invading DNA in real\u2010time has not yet been studied at the single\u2010cell level.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>In this study, we fluorescently label phage lambda DNA <jats:italic>in vivo<\/jats:italic>, and track the labeled DNA over time to characterize DNA degradation at the single\u2010cell level.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>At the bulk level, the lysogenization frequency of cells harboring CRISPR plasmids decreases significantly compared to cells with a non\u2010CRISPR control. At the single\u2010cell level, host cells with CRISPR activity are unperturbed by phage infection, maintaining normal growth like uninfected cells, where the efficiency of our anti\u2010lambda CRISPR system is around 26%. During the course of time\u2010lapse movies, the average fluorescence of invasive phage DNA in cells with CRISPR activity, decays more rapidly compared to cells without, and phage DNA is fully degraded by around 44\u00a0minutes on average. Moreover, the degradation appears to be independent of cell size or the phage DNA ejection site suggesting that Cas proteins are dispersed in sufficient quantities throughout the cell.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>With the CRISPR\u2010Cas visualization system we developed, we are able to examine and characterize how a CRISPR system degrades invading phage DNA at the single\u2010cell level. This work provides direct evidence and improves the current understanding on how CRISPR breaks down invading DNA.<\/jats:p><\/jats:sec>","DOI":"10.1007\/s40484-017-0099-0","type":"journal-article","created":{"date-parts":[[2017,3,17]],"date-time":"2017-03-17T09:05:59Z","timestamp":1489741559000},"page":"67-75","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Visualization of phage DNA degradation by a type I CRISPR\u2010Cas system at the single\u2010cell level"],"prefix":"10.1002","volume":"5","author":[{"given":"Jingwen","family":"Guan","sequence":"first","affiliation":[{"name":"<!--1--> Department of Biochemistry and Biophysics Texas A&amp;M University College Station TX 77843 USA"},{"name":"<!--2--> Center for Phage Technology Texas A&amp;M University College Station TX 77843 USA"},{"name":"<!--3--> Molecular and Environmental Plant Sciences Texas A&amp;M University College Station TX 77843 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xu","family":"Shi","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Biochemistry and Biophysics Texas A&amp;M University College Station TX 77843 USA"},{"name":"<!--2--> Center for Phage Technology Texas A&amp;M University College Station TX 77843 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Roberto","family":"Burgos","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Biochemistry and Biophysics Texas A&amp;M University College Station TX 77843 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lanying","family":"Zeng","sequence":"additional","affiliation":[{"name":"<!--1--> Department of Biochemistry and Biophysics Texas A&amp;M University College Station TX 77843 USA"},{"name":"<!--2--> Center for Phage Technology Texas A&amp;M University College Station TX 77843 USA"},{"name":"<!--3--> Molecular and Environmental Plant Sciences Texas A&amp;M University College Station TX 77843 USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2017,3]]},"reference":[{"key":"e_1_2_8_2_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.aad5147"},{"key":"e_1_2_8_3_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.micro.112408.134123"},{"key":"e_1_2_8_4_2","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkv793"},{"key":"e_1_2_8_5_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev\u2010genet\u2010110410\u2010132430"},{"key":"e_1_2_8_6_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2016.01.030"},{"key":"e_1_2_8_7_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro3279"},{"key":"e_1_2_8_8_2","doi-asserted-by":"publisher","DOI":"10.1038\/nsmb.2875"},{"key":"e_1_2_8_9_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1112832108"},{"key":"e_1_2_8_10_2","doi-asserted-by":"publisher","DOI":"10.1038\/emboj.2011.41"},{"key":"e_1_2_8_11_2","doi-asserted-by":"publisher","DOI":"10.1186\/s13059\u2010015\u20100816\u20109"},{"key":"e_1_2_8_12_2","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M113.472233"},{"key":"e_1_2_8_13_2","doi-asserted-by":"publisher","DOI":"10.1038\/nrmicro.2015.14"},{"key":"e_1_2_8_14_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2016.07.011"},{"key":"e_1_2_8_15_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2016.08.038"},{"key":"e_1_2_8_16_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1256328"},{"key":"e_1_2_8_17_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1405079111"},{"key":"e_1_2_8_18_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2012.03.018"},{"key":"e_1_2_8_19_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1256996"},{"key":"e_1_2_8_20_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2015.10.003"},{"key":"e_1_2_8_21_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1159689"},{"key":"e_1_2_8_22_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00644\u201010"},{"key":"e_1_2_8_23_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.1153498"},{"key":"e_1_2_8_24_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2015.03.027"},{"key":"e_1_2_8_25_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365\u20102958.2010.07073.x"},{"key":"e_1_2_8_26_2","doi-asserted-by":"publisher","DOI":"10.1016\/0092\u20108674(94)90156\u20102"},{"key":"e_1_2_8_27_2","doi-asserted-by":"publisher","DOI":"10.1016\/0092\u20108674(95)90272\u20104"},{"key":"e_1_2_8_28_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365\u20102958.2010.07265.x"},{"key":"e_1_2_8_29_2","doi-asserted-by":"publisher","DOI":"10.1002\/mbo3.395"},{"key":"e_1_2_8_30_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cub.2012.05.023"},{"key":"e_1_2_8_31_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365\u20102958.2008.06205.x"},{"key":"e_1_2_8_32_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.bpj.2011.05.014"},{"key":"e_1_2_8_33_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2010.03.034"},{"key":"e_1_2_8_34_2","doi-asserted-by":"publisher","DOI":"10.3791\/3363"},{"key":"e_1_2_8_35_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365\u20102958.2011.07579.x"}],"container-title":["Quantitative Biology"],"original-title":[],"language":"en","link":[{"URL":"http:\/\/link.springer.com\/article\/10.1007\/s40484-017-0099-0\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"http:\/\/link.springer.com\/content\/pdf\/10.1007\/s40484-017-0099-0.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"http:\/\/link.springer.com\/content\/pdf\/10.1007\/s40484-017-0099-0.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1007\/s40484-017-0099-0","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,1,3]],"date-time":"2025-01-03T09:58:27Z","timestamp":1735898307000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1007\/s40484-017-0099-0"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2017,3]]},"references-count":34,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2017,3]]}},"alternative-id":["10.1007\/s40484-017-0099-0"],"URL":"https:\/\/doi.org\/10.1007\/s40484-017-0099-0","archive":["Portico"],"relation":{},"ISSN":["2095-4689","2095-4697"],"issn-type":[{"value":"2095-4689","type":"print"},{"value":"2095-4697","type":"electronic"}],"subject":[],"published":{"date-parts":[[2017,3]]},"assertion":[{"value":"20 September 2016","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"22 December 2016","order":2,"name":"revised","label":"Revised","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"4 February 2017","order":3,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"30 March 2017","order":4,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}}]}}