{"doi":"10.1101/2023.02.07.527484","title":"IVT generation of guideRNAs for Cas9-enrichment Nanopore Sequencing","abstract":"ABSTRACT Generating high-coverage sequencing coverage at select genomic loci has extensive applications in both research science and genetic medicine. Long-read sequencing technologies (e.g. nanopore sequencing) have expanded our ability to generate sequencing data in regions (e.g. repetitive elements) that are difficult to interrogate with short-read sequencing methods. In work presented here, we expand on our previous work using CRISPR/Cas9 for targeted nanopore sequencing by using in vitro transcribed guideRNAs, with 1100 guideRNAs in a single experiment. This approach decreases the cost per guideRNA, increases the number of guideRNAs that can be multiplexed in a single experiment, and provides a way to rapidly screen numerous guideRNAs for cutting efficiency. We apply this strategy in multiple patient-derived pancreatic cancer cell lines, demonstrating its ability to unveil structural variation in “deletion hotspots” around the tumor suppressor genes p16 ( CDKN2A ), and SMAD4 .","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2023,"id":390348,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":11,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9512,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1162123,"name":"Josh Zhiyong Wang","orcid":null,"position":1,"is_corresponding":false},{"id":44709,"name":"David S. Weiss","orcid":"0000-0003-0980-7866","position":2,"is_corresponding":false},{"id":1161695,"name":"Alexis L. Norris","orcid":"0000-0002-0442-9096","position":3,"is_corresponding":false},{"id":62913,"name":"James R. Eshleman","orcid":"0000-0002-9274-8459","position":4,"is_corresponding":false},{"id":49049,"name":"Winston Timp","orcid":"0000-0003-2083-6027","position":5,"is_corresponding":false},{"id":505809,"name":"Timothy Gilpatrick","orcid":null,"position":0,"is_corresponding":true}],"reference_count":16,"raw_metadata":null,"created_at":"2026-07-19T01:18:32.854511Z","pmid":"36798399","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}