{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,24]],"date-time":"2026-10-24T02:19:21Z","timestamp":1792808361105,"version":"4.5.0"},"posted":{"date-parts":[[2026,9,14]]},"group-title":"Molecular Biology","reference-count":60,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,9,14]],"date-time":"2026-09-14T00:00:00Z","timestamp":1789344000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Howard Hughes Medical Institute","id":[{"id":"https:\/\/ror.org\/006w34k90","id-type":"ROR","asserted-by":"crossref"}]},{"name":"U.S. Department of Defense","award":["MD190071"],"award-info":[{"award-number":["MD190071"]}],"id":[{"id":"https:\/\/ror.org\/0447fe631","id-type":"ROR","asserted-by":"crossref"}]},{"name":"NIH \/ NINDS","award":["1R01 NS123160-01"],"award-info":[{"award-number":["1R01 NS123160-01"]}]},{"name":"Usher 1F Foundation"},{"name":"Salk Institute Gene Therapy Fund"},{"name":"W. M. Keck Foundation","id":[{"id":"https:\/\/ror.org\/000dswa46","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Mathers Foundation","id":[{"id":"https:\/\/ror.org\/02a7hjv13","id-type":"ROR","asserted-by":"crossref"}]},{"name":"Sol Goldman Charitable Trust","id":[{"id":"https:\/\/ror.org\/04w7c5n31","id-type":"ROR","asserted-by":"crossref"}]},{"name":"NIH \/ NICHD","award":["F30 HD106732"],"award-info":[{"award-number":["F30 HD106732"]}]},{"name":"NIH \/ NCI"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,9,14]]},"abstract":"<jats:title>Summary<\/jats:title>\n                <jats:p>Gene complementation is a powerful tool for genetic selection and protein functional studies but typically requires extensive screening for complementary components. Here we engineered a platform for gene complementation based on RNA end-joining (REJ) that precisely and efficiently splices separate RNA units into functional coding mRNAs. REJ is mediated by short structured modular RNA segments of \u223c250bp that: (a) promote RNA::RNA interaction, (b) coopt the intrinsic cell splicing machinery to facilitate RNA trans-splicing, (c) minimizes translation of protein fragments from un-spliced RNA segments, and (d) encodes scar-free protein. We demonstrate that REJ is a broadly applicable system that can reliably split nearly any gene into complementary segments regardless of protein structure. To enable the use of this system we provide a toolbox of reporters and a web-based design tool to facilitate vector design. Demonstrated REJ applications include genetic complementation, intersectional labeling of cell types, and efficient expression of large proteins.<\/jats:p>\n                <jats:sec>\n                  <jats:title>Highlights<\/jats:title>\n                  <jats:list list-type=\"bullet\">\n                    <jats:list-item>\n                      <jats:p>\n                        Synthetic RNA cis-elements can direct efficient and precise RNA trans-splicing\n                        <jats:italic>in vivo<\/jats:italic>\n                        .\n                      <\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Endogenous splicing factors catalyze RNA end-joining (REJ) without foreign proteins.<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>The REJ platform can be applied to nearly any gene with few positional constraints.<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>REJ can be used as an intersectional tool to build complex genetic logic circuits.<\/jats:p>\n                    <\/jats:list-item>\n                  <\/jats:list>\n                <\/jats:sec>","DOI":"10.64898\/2026.09.13.751279","type":"posted-content","created":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T05:00:22Z","timestamp":1789448422000},"source":"Crossref","is-referenced-by-count":0,"title":["An intersectional expression platform for gene complementation using RNA-fragment end joining (REJ)"],"prefix":"10.64898","author":[{"given":"Lukas C.","family":"Bachmann","sequence":"first","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"},{"name":"Insmed, San Diego. 9330 Scranton Road Suite 200, San Diego, CA 92121, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8221-224X","authenticated-orcid":false,"given":"Ryan H.","family":"Hsu","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"},{"name":"Bioengineering, Jacobs School of Engineering, University of California, San Diego, La Jolla, CA 92092, USA"},{"name":"Medical Scientist Training Program, School of Medicine, University of California, San Diego, La Jolla 92092, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kip J.","family":"Hermann","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Claire E.","family":"Williams","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard L.","family":"Farman","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicolas","family":"Criales","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Craig","family":"Clark","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stella","family":"Kramer","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carolina Th\u00f6rn","family":"Perez","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sawyer","family":"Randles","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"},{"name":"Biological Sciences Graduate Program, University of California San Diego, La Jolla, CA 92092, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karen","family":"Lettieri","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Samuel L.","family":"Pfaff","sequence":"additional","affiliation":[{"name":"Gene Expression Laboratory, Salk Institute for Biological Studies, 10010 North Torrey Pines, La Jolla, CA 92037, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026091707201263000_2026.09.13.751279v1.1","doi-asserted-by":"publisher","DOI":"10.1016\/0167-7799(93)90069-L"},{"key":"2026091707201263000_2026.09.13.751279v1.2","doi-asserted-by":"publisher","DOI":"10.1093\/hmg\/ddu125"},{"key":"2026091707201263000_2026.09.13.751279v1.3","doi-asserted-by":"publisher","DOI":"10.2174\/156652321402140318165450"},{"key":"2026091707201263000_2026.09.13.751279v1.4","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-020-1978-5"},{"key":"2026091707201263000_2026.09.13.751279v1.5","doi-asserted-by":"publisher","DOI":"10.1016\/j.mayocp.2023.11.003"},{"key":"2026091707201263000_2026.09.13.751279v1.6","doi-asserted-by":"publisher","DOI":"10.1517\/14728222.11.6.835"},{"key":"2026091707201263000_2026.09.13.751279v1.7","doi-asserted-by":"publisher","DOI":"10.1038\/nsb887"},{"key":"2026091707201263000_2026.09.13.751279v1.8","doi-asserted-by":"publisher","DOI":"10.1101\/cshperspect.a003707"},{"key":"2026091707201263000_2026.09.13.751279v1.9","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2019.09.017"},{"key":"2026091707201263000_2026.09.13.751279v1.10","doi-asserted-by":"publisher","DOI":"10.1038\/nature07509"},{"key":"2026091707201263000_2026.09.13.751279v1.11","doi-asserted-by":"publisher","DOI":"10.1016\/0168-9525(88)90107-2"},{"key":"2026091707201263000_2026.09.13.751279v1.12","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-024-46172-4"},{"key":"2026091707201263000_2026.09.13.751279v1.13","doi-asserted-by":"publisher","DOI":"10.1038\/6986"},{"key":"2026091707201263000_2026.09.13.751279v1.14","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(87)90207-8"},{"key":"2026091707201263000_2026.09.13.751279v1.15","doi-asserted-by":"publisher","DOI":"10.1038\/jid.2012.101"},{"key":"2026091707201263000_2026.09.13.751279v1.16","doi-asserted-by":"publisher","DOI":"10.1002\/ctm2.1607"},{"key":"2026091707201263000_2026.09.13.751279v1.17","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-4939-3271-9_2"},{"key":"2026091707201263000_2026.09.13.751279v1.18","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-023-42386-0"},{"key":"2026091707201263000_2026.09.13.751279v1.19","doi-asserted-by":"publisher","DOI":"10.1002\/emmm.201302948"},{"key":"2026091707201263000_2026.09.13.751279v1.20","doi-asserted-by":"publisher","DOI":"10.1016\/S0955-0674(97)80007-5"},{"key":"2026091707201263000_2026.09.13.751279v1.21","doi-asserted-by":"publisher","DOI":"10.1089\/hum.2008.173"},{"key":"2026091707201263000_2026.09.13.751279v1.22","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymthe.2003.08.022"},{"key":"2026091707201263000_2026.09.13.751279v1.23","first-page":"141","article-title":"Spliceosome-mediated RNA trans-splicing in gene therapy and genomics","volume":"1","year":"2001","journal-title":"Gene Therapy and Regulation"},{"key":"2026091707201263000_2026.09.13.751279v1.24","doi-asserted-by":"publisher","DOI":"10.1006\/jmbi.1996.0301"},{"key":"2026091707201263000_2026.09.13.751279v1.25","doi-asserted-by":"publisher","DOI":"10.1016\/S0968-0004(03)00052-5"},{"key":"2026091707201263000_2026.09.13.751279v1.26","doi-asserted-by":"publisher","DOI":"10.1007\/978-3-540-34449-0_4"},{"key":"2026091707201263000_2026.09.13.751279v1.27","doi-asserted-by":"publisher","DOI":"10.1007\/978-0-387-77374-2_6"},{"key":"2026091707201263000_2026.09.13.751279v1.28","doi-asserted-by":"publisher","DOI":"10.1080\/15476286.2016.1166330"},{"key":"2026091707201263000_2026.09.13.751279v1.29","doi-asserted-by":"publisher","DOI":"10.1101\/gad.1163204"},{"key":"2026091707201263000_2026.09.13.751279v1.30","doi-asserted-by":"publisher","DOI":"10.1016\/j.biocel.2017.06.016"},{"key":"2026091707201263000_2026.09.13.751279v1.31","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-4939-9139-6_2"},{"key":"2026091707201263000_2026.09.13.751279v1.32","doi-asserted-by":"publisher","DOI":"10.1089\/hgtb.2015.140"},{"key":"2026091707201263000_2026.09.13.751279v1.33","doi-asserted-by":"publisher","DOI":"10.1186\/s13578-024-01272-y"},{"key":"2026091707201263000_2026.09.13.751279v1.34","doi-asserted-by":"publisher","DOI":"10.1089\/hum.2023.017"},{"key":"2026091707201263000_2026.09.13.751279v1.35","doi-asserted-by":"publisher","DOI":"10.1007\/978-1-4939-7522-8_11"},{"key":"2026091707201263000_2026.09.13.751279v1.36","doi-asserted-by":"publisher","DOI":"10.1126\/science.adp8179"},{"key":"2026091707201263000_2026.09.13.751279v1.37","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2025.12.039"},{"key":"2026091707201263000_2026.09.13.751279v1.38","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-immunol-042718-041356"},{"key":"2026091707201263000_2026.09.13.751279v1.39","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2006.02.015"},{"key":"2026091707201263000_2026.09.13.751279v1.40","doi-asserted-by":"publisher","DOI":"10.1038\/s41392-021-00687-0"},{"key":"2026091707201263000_2026.09.13.751279v1.41","doi-asserted-by":"publisher","DOI":"10.3389\/fnana.2019.00093"},{"key":"2026091707201263000_2026.09.13.751279v1.42","doi-asserted-by":"publisher","DOI":"10.1016\/j.tibtech.2005.03.006"},{"key":"2026091707201263000_2026.09.13.751279v1.43","doi-asserted-by":"publisher","DOI":"10.1126\/scitranslmed.aav4523"},{"key":"2026091707201263000_2026.09.13.751279v1.44","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-024-07710-8"},{"key":"2026091707201263000_2026.09.13.751279v1.45","doi-asserted-by":"publisher","DOI":"10.1006\/mthe.2001.0456"},{"key":"2026091707201263000_2026.09.13.751279v1.46","doi-asserted-by":"publisher","DOI":"10.1105\/tpc.107.055053"},{"key":"2026091707201263000_2026.09.13.751279v1.47","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymthe.2023.10.015"},{"key":"2026091707201263000_2026.09.13.751279v1.48","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2023.11.033"},{"key":"2026091707201263000_2026.09.13.751279v1.49","doi-asserted-by":"publisher","DOI":"10.1261\/rna.079016.121"},{"key":"2026091707201263000_2026.09.13.751279v1.50","doi-asserted-by":"publisher","DOI":"10.1038\/s41598-023-33208-w"},{"key":"2026091707201263000_2026.09.13.751279v1.51","doi-asserted-by":"publisher","DOI":"10.1016\/j.immuni.2020.03.017"},{"key":"2026091707201263000_2026.09.13.751279v1.52","doi-asserted-by":"publisher","DOI":"10.1016\/j.coviro.2015.01.011"},{"key":"2026091707201263000_2026.09.13.751279v1.53","doi-asserted-by":"publisher","DOI":"10.1101\/2021.01.09.426034"},{"key":"2026091707201263000_2026.09.13.751279v1.54","doi-asserted-by":"publisher","DOI":"10.1016\/j.jtbi.2020.110336"},{"key":"2026091707201263000_2026.09.13.751279v1.55","first-page":"611","article-title":"Adeno-associated Virus (AAV) Dual Vector Strategies for Gene Therapy Encoding Large Transgenes","volume":"90","year":"2017","journal-title":"Yale J Biol Med"},{"key":"2026091707201263000_2026.09.13.751279v1.56","doi-asserted-by":"publisher","DOI":"10.1089\/hum.2008.009"},{"key":"2026091707201263000_2026.09.13.751279v1.57","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.161269798"},{"key":"2026091707201263000_2026.09.13.751279v1.58","doi-asserted-by":"publisher","DOI":"10.3390\/genes10040287"},{"key":"2026091707201263000_2026.09.13.751279v1.59","doi-asserted-by":"publisher","DOI":"10.1016\/j.ymthe.2017.03.013"},{"key":"2026091707201263000_2026.09.13.751279v1.60","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.bi.44.070175.001455"}],"container-title":[],"original-title":[],"link":[{"URL":"https:\/\/syndication.highwire.org\/content\/doi\/10.64898\/2026.09.13.751279","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,17]],"date-time":"2026-09-17T14:20:22Z","timestamp":1789654822000},"score":1,"resource":{"primary":{"URL":"http:\/\/biorxiv.org\/lookup\/doi\/10.64898\/2026.09.13.751279"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,9,14]]},"references-count":60,"URL":"https:\/\/doi.org\/10.64898\/2026.09.13.751279","relation":{},"subject":[],"published":{"date-parts":[[2026,9,14]]},"subtype":"preprint"}}