{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T23:07:02Z","timestamp":1788995222313,"version":"build-2803163510"},"reference-count":27,"publisher":"Oxford University Press (OUP)","issue":"5","license":[{"start":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T00:00:00Z","timestamp":1788912000000},"content-version":"vor","delay-in-days":8,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["32572957"],"award-info":[{"award-number":["32572957"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,9,1]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>The precision of CRISPR\/Cas systems is fundamental to their application in plant and animal biotechnology. However, comprehensive sequence-based off-target candidate discovery remains a computational bottleneck, particularly in large and complex genomes. Here we developed SpacerScope, an off-target candidate discovery framework that enables unbiased, genome-wide discovery by leveraging binary vectorization, bitwise filtering, and right-end-anchored alignment. Benchmarking against human CIRCLE-seq data demonstrated that SpacerScope recovered 100% of validated off-target sites (6142\/6142), matching the sensitivity of exhaustive algorithms. Crucially, SpacerScope achieved this maximum candidate recovery while substantially reducing computational overhead. In large-genome evaluations, SpacerScope maintained low peak memory usage of 2.20 GiB and achieved substantial runtime improvements over indel-aware comparator tools, including more than 50-fold speedup relative to Cas-OFFinder 3 (544\u00a0s versus 29\u00a0185\u00a0s). Furthermore, comparative analyses in polyploid species, such as the octoploid strawberry, revealed that SpacerScope identified larger sequence-compatible candidate burdens than standard web-based design platforms. Our results establish SpacerScope as a high-speed framework for sequence-based genome-wide off-target candidate discovery across diverse and highly repetitive genomic landscapes. The source code and program was publicly available at https:\/\/github.com\/charlesqu666\/SpacerScope.<\/jats:p>\n                  <jats:p>Short Abstract CRISPR\/Cas sequence-based off-target candidate discovery remains computationally challenging in large, repetitive, and polyploid genomes. Existing tools either miss indel-containing candidate sites or incur prohibitive runtime and memory costs. We developed SpacerScope, a binary-vectorized framework that enables unbiased, genome-wide off-target candidate discovery without pre-selected candidate sites. By integrating bitwise filtering with right-end-anchored alignment, SpacerScope recovered 100% of validated off-target sites in human CIRCLE-seq data while using only 2.20 GiB of memory and achieving more than 10-fold speedup over indel-aware alternatives. Evaluation in plant genomes, including rice and octoploid strawberry, further demonstrated SpacerScope\u2019s capacity to identify larger sequence-compatible candidate burdens overlooked by standard tools. SpacerScope thus provides a high-speed framework for sequence-based genome-wide off-target candidate discovery across diverse and highly repetitive genomic landscapes, supporting downstream prioritization.<\/jats:p>","DOI":"10.1093\/bib\/bbag478","type":"journal-article","created":{"date-parts":[[2026,8,14]],"date-time":"2026-08-14T11:14:46Z","timestamp":1786706086000},"source":"Crossref","is-referenced-by-count":0,"title":["SpacerScope: binary-vectorized, genome-wide off-target profiling for RNA-guided nucleases without prior candidate-site bias"],"prefix":"10.1093","volume":"27","author":[{"given":"Yanji","family":"Qu","sequence":"first","affiliation":[{"name":"College of Horticulture, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yaxuan","family":"Wang","sequence":"additional","affiliation":[{"name":"College of Horticulture, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yan","family":"Wang","sequence":"additional","affiliation":[{"name":"College of Horticulture, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haoru","family":"Tang","sequence":"additional","affiliation":[{"name":"College of Horticulture, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province,","place":["China"]},{"name":"Key Laboratory of Agricultural Bioinformatics, Ministry of Education, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1276-1064","authenticated-orcid":false,"given":"Qing","family":"Chen","sequence":"additional","affiliation":[{"name":"College of Horticulture, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province,","place":["China"]},{"name":"Key Laboratory of Agricultural Bioinformatics, Ministry of Education, Sichuan Agricultural University , Huimin Road 211, Wenjiang District, Chengdu City 611130, Sichuan Province ,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2026,9,9]]},"reference":[{"key":"2026090918202158100_ref1","doi-asserted-by":"publisher","first-page":"1473","DOI":"10.1093\/bioinformatics\/btu048","article-title":"Cas-OFFinder: a fast and versatile algorithm that searches for potential off-target sites of Cas9 RNA-guided 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