{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T21:51:28Z","timestamp":1791323488848,"version":"4.3.1"},"reference-count":37,"publisher":"Elsevier BV","license":[{"start":{"date-parts":[[2026,12,1]],"date-time":"2026-12-01T00:00:00Z","timestamp":1796083200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/www.elsevier.com\/tdm\/userlicense\/1.0\/"},{"start":{"date-parts":[[2026,12,1]],"date-time":"2026-12-01T00:00:00Z","timestamp":1796083200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/www.elsevier.com\/legal\/tdmrep-license"},{"start":{"date-parts":[[2026,9,24]],"date-time":"2026-09-24T00:00:00Z","timestamp":1790208000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100002322","name":"Coordena\u00e7\u00e3o de Aperfei\u00e7oamento de Pessoal de N\u00edvel Superior","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100002322","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003593","name":"Conselho Nacional de Desenvolvimento Cient\u00edfico e Tecnol\u00f3gico","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100003593","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100006182","name":"FAPES","doi-asserted-by":"publisher","award":["T.O. 877\/2023 \\u2212 P:2023-QQ9QC Edital FAPES N. 28\/2022 \\u2212 Universal"],"award-info":[{"award-number":["T.O. 877\/2023 \\u2212 P:2023-QQ9QC Edital FAPES N. 28\/2022 \\u2212 Universal"]}],"id":[{"id":"10.13039\/501100006182","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["elsevier.com","sciencedirect.com"],"crossmark-restriction":true},"short-container-title":["Methods"],"published-print":{"date-parts":[[2026,12]]},"DOI":"10.1016\/j.ymeth.2026.09.004","type":"journal-article","created":{"date-parts":[[2026,9,26]],"date-time":"2026-09-26T15:03:15Z","timestamp":1790434995000},"page":"102-108","update-policy":"https:\/\/doi.org\/10.1016\/elsevier_cm_policy","source":"Crossref","is-referenced-by-count":0,"special_numbering":"C","title":["In vitro validation of a CRISPR\/Cas9 guide RNA targeting COL1A1 gene by cleavage assay"],"prefix":"10.1016","volume":"256","author":[{"given":"Daniel Moreira","family":"Melo","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sabrina Rodrigues","family":"Sarquis","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paolla Manzoli","family":"Luz","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Raphael Cesar","family":"Barcellos Bevitori","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yasmin","family":"de Almeida Barbosa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Roger Frossard","family":"Pagotto","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maira","family":"Trancozo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eldam\u00e1ria","family":"de Vargas Wolfgramm dos Santos","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Flavia","family":"de Paula","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"78","reference":[{"issue":"6176","key":"10.1016\/j.ymeth.2026.09.004_b0005","doi-asserted-by":"crossref","DOI":"10.1126\/science.1247997","article-title":"Structures of Cas9 endonucleases reveal RNA-mediated conformational activation","volume":"343","author":"Jinek","year":"2014","journal-title":"Science"},{"issue":"6213","key":"10.1016\/j.ymeth.2026.09.004_b0010","doi-asserted-by":"crossref","DOI":"10.1126\/science.1258096","article-title":"The new frontier of genome engineering with CRISPR-Cas9","volume":"346","author":"Doudna","year":"2014","journal-title":"Science"},{"issue":"19","key":"10.1016\/j.ymeth.2026.09.004_b0015","doi-asserted-by":"crossref","first-page":"10355","DOI":"10.3390\/ijms221910355","article-title":"Comparison of the feasibility, efficiency, and safety of genome editing technologies","volume":"22","author":"Gonz\u00e1lez Castro","year":"2021","journal-title":"Int. J. Mol. Sci."},{"issue":"1","key":"10.1016\/j.ymeth.2026.09.004_b0020","doi-asserted-by":"crossref","first-page":"319","DOI":"10.3390\/ijms22010319","article-title":"New strategies to overcome present CRISPR\/Cas9 limitations in apple and pear: efficient dechimerization and base editing","volume":"22","author":"Malabarba","year":"2020","journal-title":"Int. J. Mol. Sci."},{"issue":"5","key":"10.1016\/j.ymeth.2026.09.004_b0025","doi-asserted-by":"crossref","first-page":"4879","DOI":"10.1007\/s11033-021-06479-7","article-title":"CRISPR\/Cas based gene editing: marking a new era in medical science","volume":"48","author":"Prasad","year":"2021","journal-title":"Mol. Biol. Rep."},{"issue":"9","key":"10.1016\/j.ymeth.2026.09.004_b0030","doi-asserted-by":"crossref","DOI":"10.1002\/pld3.269","article-title":"Versatile in vitro assay to recognize Cas9\u2010induced mutations","volume":"4","author":"Bente","year":"2020","journal-title":"Plant Direct"},{"key":"10.1016\/j.ymeth.2026.09.004_b0035","doi-asserted-by":"crossref","unstructured":"S. Karmakar, D. Behera, M.J. Baig, K.A. Molla. (2021). In vitro Cas9 Cleavage Assay to Check Guide RNA Efficiency (pp. 23\u201339). https:\/\/doi.org\/10.1007\/978-1-0716-1657-4_3.","DOI":"10.1007\/978-1-0716-1657-4_3"},{"issue":"6","key":"10.1016\/j.ymeth.2026.09.004_b0040","doi-asserted-by":"crossref","first-page":"891","DOI":"10.1007\/s00223-024-01266-5","article-title":"Update on the genetics of osteogenesis imperfecta","volume":"115","author":"Jovanovic","year":"2024","journal-title":"Calcif. Tissue Int."},{"issue":"3","key":"10.1016\/j.ymeth.2026.09.004_b0045","doi-asserted-by":"crossref","first-page":"209","DOI":"10.1002\/humu.20429","article-title":"Consortium for osteogenesis imperfecta mutations in the helical domain of type I collagen: regions rich in lethal mutations align with collagen binding sites for integrins and proteoglycans","volume":"28","author":"Marini","year":"2007","journal-title":"Hum. Mutat."},{"issue":"4","key":"10.1016\/j.ymeth.2026.09.004_b0050","doi-asserted-by":"crossref","first-page":"R95","DOI":"10.1530\/EJE-20-0299","article-title":"Management of endocrine disease: osteogenesis imperfecta: an update on clinical features and therapies","volume":"183","author":"Marom","year":"2020","journal-title":"Eur. J. Endocrinol."},{"key":"10.1016\/j.ymeth.2026.09.004_b0055","doi-asserted-by":"crossref","DOI":"10.1016\/j.scr.2019.101453","article-title":"The use of simultaneous reprogramming and gene correction to generate an osteogenesis imperfecta patient COL1A1 c. 3936 G&gt;T iPSC line and an isogenic control iPSC line","volume":"38","author":"Howden","year":"2019","journal-title":"Stem Cell Res."},{"issue":"14","key":"10.1016\/j.ymeth.2026.09.004_b0060","doi-asserted-by":"crossref","first-page":"3141","DOI":"10.3390\/jcm10143141","article-title":"Restoration of osteogenesis by CRISPR\/Cas9 genome editing of the mutated COL1A1 gene in osteogenesis imperfecta","volume":"10","author":"Jung","year":"2021","journal-title":"J. Clin. Med."},{"issue":"3","key":"10.1016\/j.ymeth.2026.09.004_b0065","doi-asserted-by":"crossref","first-page":"817","DOI":"10.1007\/s13577-024-01028-3","article-title":"Identification of a family with van der Hoeve\u2019s syndrome harboring a novel COL1A1 mutation and generation of patient-derived iPSC lines and CRISPR\/Cas9-corrected isogenic iPSCs","volume":"37","author":"Li","year":"2024","journal-title":"Hum. Cell"},{"key":"10.1016\/j.ymeth.2026.09.004_b0070","doi-asserted-by":"crossref","first-page":"646","DOI":"10.1016\/j.bone.2019.07.021","article-title":"A novel transgenic murine model with persistently brittle bones simulating osteogenesis imperfecta type I","volume":"127","author":"Liu","year":"2019","journal-title":"Bone"},{"key":"10.1016\/j.ymeth.2026.09.004_b0075","doi-asserted-by":"crossref","unstructured":"K. Labun, O. Rio, H. Tjeldnes, M. Swirski, A.Z. Komisarczuk, E. Haapaniemi, E. Valen. (2025). CHOPOFF: symbolic alignments enable fast and sensitive CRISPR off-target detection. https:\/\/doi.org\/10.1101\/2025.01.06.603201.","DOI":"10.1101\/2025.01.06.603201"},{"issue":"2","key":"10.1016\/j.ymeth.2026.09.004_b0080","doi-asserted-by":"crossref","first-page":"211","DOI":"10.1089\/107632701300062859","article-title":"Multilineage cells from human adipose tissue: implications for cell-based therapies","volume":"7","author":"Zuk","year":"2001","journal-title":"Tissue Eng."},{"issue":"4","key":"10.1016\/j.ymeth.2026.09.004_b0085","doi-asserted-by":"crossref","first-page":"287","DOI":"10.4161\/org.27153","article-title":"A rapid and efficient method for primary culture of human adipose-derived stem cells","volume":"9","author":"Zeng","year":"2013","journal-title":"Organogenesis"},{"issue":"3","key":"10.1016\/j.ymeth.2026.09.004_b0090","doi-asserted-by":"crossref","first-page":"1215","DOI":"10.1093\/nar\/16.3.1215","article-title":"A simple salting out procedure for extracting DNA from human nucleated cells","volume":"16","author":"Miller","year":"1988","journal-title":"Nucleic Acids Res."},{"issue":"12","key":"10.1016\/j.ymeth.2026.09.004_b0095","doi-asserted-by":"crossref","DOI":"10.1371\/journal.pone.0226107","article-title":"Optimizing sgRNA length to improve target specificity and efficiency for the GGTA1 gene using the CRISPR\/Cas9 gene editing system","volume":"14","author":"Matson","year":"2019","journal-title":"PLoS One"},{"issue":"W1","key":"10.1016\/j.ymeth.2026.09.004_b0100","doi-asserted-by":"crossref","first-page":"W171","DOI":"10.1093\/nar\/gkz365","article-title":"CHOPCHOP v3: expanding the CRISPR web toolbox beyond genome editing","volume":"47","author":"Labun","year":"2019","journal-title":"Nucleic Acids Res."},{"issue":"8","key":"10.1016\/j.ymeth.2026.09.004_b0105","doi-asserted-by":"crossref","first-page":"1147","DOI":"10.1101\/gr.191452.115","article-title":"Sequence determinants of improved CRISPR sgRNA design","volume":"25","author":"Xu","year":"2015","journal-title":"Genome Res."},{"issue":"2","key":"10.1016\/j.ymeth.2026.09.004_b0110","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1038\/nbt.3437","article-title":"Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9","volume":"34","author":"Doench","year":"2016","journal-title":"Nat. Biotechnol."},{"issue":"9","key":"10.1016\/j.ymeth.2026.09.004_b0115","doi-asserted-by":"crossref","first-page":"827","DOI":"10.1038\/nbt.2647","article-title":"DNA targeting specificity of RNA-guided Cas9 nucleases","volume":"31","author":"Hsu","year":"2013","journal-title":"Nat. Biotechnol."},{"issue":"7519","key":"10.1016\/j.ymeth.2026.09.004_b0120","doi-asserted-by":"crossref","first-page":"569","DOI":"10.1038\/nature13579","article-title":"Structural basis of PAM-dependent target DNA recognition by the Cas9 endonuclease","volume":"513","author":"Anders","year":"2014","journal-title":"Nature"},{"issue":"9","key":"10.1016\/j.ymeth.2026.09.004_b0125","doi-asserted-by":"crossref","first-page":"839","DOI":"10.1038\/nbt.2673","article-title":"High-throughput profiling of off-target DNA cleavage reveals RNA-programmed Cas9 nuclease specificity","volume":"31","author":"Pattanayak","year":"2013","journal-title":"Nat. Biotechnol."},{"issue":"1","key":"10.1016\/j.ymeth.2026.09.004_b0130","doi-asserted-by":"crossref","first-page":"40638","DOI":"10.1038\/srep40638","article-title":"Profiling single-guide RNA specificity reveals a mismatch sensitive core sequence","volume":"7","author":"Zheng","year":"2017","journal-title":"Sci. Rep."},{"issue":"22","key":"10.1016\/j.ymeth.2026.09.004_b0135","doi-asserted-by":"crossref","first-page":"4067","DOI":"10.1016\/j.cell.2022.09.026","article-title":"Structural basis for Cas9 off-target activity","volume":"185","author":"Pacesa","year":"2022","journal-title":"Cell"},{"issue":"7","key":"10.1016\/j.ymeth.2026.09.004_b0140","doi-asserted-by":"crossref","first-page":"677","DOI":"10.1038\/nbt.2916","article-title":"Genome-wide analysis reveals characteristics of off-target sites bound by the Cas9 endonuclease","volume":"32","author":"Kuscu","year":"2014","journal-title":"Nat. Biotechnol."},{"issue":"5","key":"10.1016\/j.ymeth.2026.09.004_b0145","doi-asserted-by":"crossref","first-page":"239","DOI":"10.1016\/j.jgg.2016.04.008","article-title":"Genome editing with CRISPR-Cas9: can it get any better?","volume":"43","author":"Haeussler","year":"2016","journal-title":"J. Genet. Genomics"},{"issue":"1","key":"10.1016\/j.ymeth.2026.09.004_b0150","doi-asserted-by":"crossref","first-page":"533","DOI":"10.1093\/molbev\/msq221","article-title":"Comparative vertebrate evolutionary analyses of type I Collagen: potential of COL1a1 gene structure and intron variation for common bone-related diseases","volume":"28","author":"Stover","year":"2011","journal-title":"Mol. Biol. Evol."},{"key":"10.1016\/j.ymeth.2026.09.004_b0155","article-title":"Optimizing PCR amplification of GC-rich nicotinic acetylcholine receptor subunits from invertebrates","volume":"42","author":"Khan","year":"2025","journal-title":"Biochem. Biophys. Rep."},{"issue":"7","key":"10.1016\/j.ymeth.2026.09.004_b0160","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1016\/j.matbio.2010.08.002","article-title":"Expression in SPARC-null mice of collagen type I lacking the globular domain of the \u03b11(I) N-propeptide results in abdominal hernias and loss of dermal collagen","volume":"29","author":"Card","year":"2010","journal-title":"Matrix Biol."},{"issue":"1","key":"10.1016\/j.ymeth.2026.09.004_b0165","doi-asserted-by":"crossref","first-page":"79","DOI":"10.18388\/abp.2017_1612","article-title":"Mutations in COL1A1 and COL1A2 genes associated with osteogenesis imperfecta (OI) types I or III","volume":"65","author":"Augusciak-Duma","year":"2018","journal-title":"Acta Biochim. Pol."},{"issue":"1","key":"10.1016\/j.ymeth.2026.09.004_b0170","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1186\/1471-2350-15-45","article-title":"COL1A1and miR-29b show lower expression levels during osteoblast differentiation of bone marrow stromal cells from osteogenesis imperfecta patients","volume":"15","author":"Kaneto","year":"2014","journal-title":"BMC Med. Genet."},{"issue":"5","key":"10.1016\/j.ymeth.2026.09.004_b0175","doi-asserted-by":"crossref","first-page":"e2213","DOI":"10.1371\/journal.pone.0002213","article-title":"Replicative senescence of mesenchymal stem cells: a continuous and organized process","volume":"3","author":"Wagner","year":"2008","journal-title":"PLoS One"},{"issue":"4","key":"10.1016\/j.ymeth.2026.09.004_b0180","doi-asserted-by":"crossref","first-page":"446","DOI":"10.3390\/antiox14040446","article-title":"Replicative senescence in mesenchymal stem cells: an in vitro study on mitochondrial dynamics and metabolic alterations","volume":"14","author":"Casorati","year":"2025","journal-title":"Antioxidants"},{"issue":"16","key":"10.1016\/j.ymeth.2026.09.004_b0185","doi-asserted-by":"crossref","first-page":"12716","DOI":"10.3390\/ijms241612716","article-title":"Practical use of immortalized cells in medicine: current advances and future perspectives","volume":"24","author":"Voloshin","year":"2023","journal-title":"Int. J. Mol. Sci."}],"container-title":["Methods"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.elsevier.com\/content\/article\/PII:S1046202326002021?httpAccept=text\/xml","content-type":"text\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/api.elsevier.com\/content\/article\/PII:S1046202326002021?httpAccept=text\/plain","content-type":"text\/plain","content-version":"vor","intended-application":"text-mining"}],"deposited":{"date-parts":[[2026,10,6]],"date-time":"2026-10-06T21:11:34Z","timestamp":1791321094000},"score":1,"resource":{"primary":{"URL":"https:\/\/linkinghub.elsevier.com\/retrieve\/pii\/S1046202326002021"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,12]]},"references-count":37,"alternative-id":["S1046202326002021"],"URL":"https:\/\/doi.org\/10.1016\/j.ymeth.2026.09.004","relation":{},"ISSN":["1046-2023"],"issn-type":[{"value":"1046-2023","type":"print"}],"subject":[],"published":{"date-parts":[[2026,12]]},"assertion":[{"value":"Elsevier","name":"publisher","label":"This article is maintained by"},{"value":"In vitro validation of a CRISPR\/Cas9 guide RNA targeting COL1A1 gene by cleavage assay","name":"articletitle","label":"Article Title"},{"value":"Methods","name":"journaltitle","label":"Journal Title"},{"value":"https:\/\/doi.org\/10.1016\/j.ymeth.2026.09.004","name":"articlelink","label":"CrossRef DOI link to publisher maintained version"},{"value":"article","name":"content_type","label":"Content Type"},{"value":"\u00a9 2026 The Author(s). Published by Elsevier Inc.","name":"copyright","label":"Copyright"}]}}