{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T18:15:31Z","timestamp":1790705731383,"version":"4.1.0"},"reference-count":29,"publisher":"Wiley","issue":"3","license":[{"start":{"date-parts":[[2024,3,13]],"date-time":"2024-03-13T00:00:00Z","timestamp":1710288000000},"content-version":"vor","delay-in-days":12,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biotechnology Journal"],"published-print":{"date-parts":[[2024,3]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>The recombinant adeno\u2010associated virus (rAAV) vectors used in gene therapy are usually produced by transfecting three different plasmids (Adenoviral helper plasmid (pHelper), AAV rep\/cap plasmids (pRepCap), and Transgene plasmid (pAAV\u2010GOI)) into human embryonic kidney 293 (HEK293) cells. However, the high proportion of unwanted empty capsids generated during rAAV production is problematic. To simultaneously enhance the genome titer and full capsid ratio, the ratio of the three plasmids transfected into HEK293 cells was optimized using design\u2010of\u2010experiment (DoE). AAV2 and AAV9, which have different production kinetics, were selected as cell\u2010associated and secreted model AAVs, respectively. In 125\u00a0mL Erlenmeyer flasks, the genome titers of rAAV2 and rAAV9 at DoE\u2010optimized plasmid weight ratios (pHelper:pRep2Cap2:pAAV\u2010GOI\u00a0=\u00a01:3.52:0.50 for rAAV2 and pHelper:pRep2Cap9:pAAV\u2010GOI\u00a0=\u00a01:1.44:0.27 for rAAV9) were 2.23\u2010fold and 2.26\u2010fold higher than those in the widely used plasmid weight ratio (1:1:1), respectively. In addition, compared with the plasmid ratio of 1:1:1, the relative VP3 band intensities of rAAV2 and rAAV9, which represent the relative empty capsid ratios, were reduced by 26% and 25%, respectively, at the DoE\u2010optimized plasmid ratio. Reduced empty capsid ratios in the DoE\u2010optimized plasmid ratios were also confirmed using transmission electron microscopy (TEM). Taken together, regardless of the AAV serotype, DoE\u2010aided optimization of the triple plasmid ratio was found to be an efficient means of improving the production of rAAV with a high full capsid ratio.<\/jats:p>","DOI":"10.1002\/biot.202300667","type":"journal-article","created":{"date-parts":[[2024,3,13]],"date-time":"2024-03-13T22:04:40Z","timestamp":1710367480000},"source":"Crossref","is-referenced-by-count":27,"title":["Enhancing the production of adeno\u2010associated virus (AAV)2 and AAV9 with high full capsid ratio in HEK293 cells through design\u2010of\u2010experiment optimization of triple plasmid ratio"],"prefix":"10.1002","volume":"19","author":[{"given":"Sungje","family":"Park","sequence":"first","affiliation":[{"name":"Department of Biological Sciences KAIST  Daejeon Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9679-1569","authenticated-orcid":false,"given":"Seunghyeon","family":"Shin","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences KAIST  Daejeon Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Haeshin","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Chemistry KAIST  Daejeon Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jae\u2010Hyung","family":"Jang","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biomolecular Engineering Yonsei University  Seoul Republic of Korea"},{"name":"R&amp;D Center GluGene Therapeutics Inc.  Seoul Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9876-7378","authenticated-orcid":false,"given":"Gyun Min","family":"Lee","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences KAIST  Daejeon Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2024,3,13]]},"reference":[{"key":"e_1_2_10_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtm.2018.11.004"},{"key":"e_1_2_10_3_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtm.2018.05.001"},{"key":"e_1_2_10_4_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41576-019-0205-4"},{"key":"e_1_2_10_5_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.202200450"},{"key":"e_1_2_10_6_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.202300051"},{"key":"e_1_2_10_7_1","doi-asserted-by":"publisher","DOI":"10.3390\/biology12020186"},{"key":"e_1_2_10_8_1","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-virology-031413-085355"},{"key":"e_1_2_10_9_1","doi-asserted-by":"publisher","DOI":"10.3390\/cells12050785"},{"key":"e_1_2_10_10_1","doi-asserted-by":"publisher","DOI":"10.1089\/hum.2010.107"},{"key":"e_1_2_10_11_1","doi-asserted-by":"publisher","DOI":"10.1089\/hum.2010.055"},{"key":"e_1_2_10_12_1","doi-asserted-by":"publisher","DOI":"10.1038\/mtm.2016.15"},{"key":"e_1_2_10_13_1","doi-asserted-by":"publisher","DOI":"10.2174\/156652310793797685"},{"key":"e_1_2_10_14_1","doi-asserted-by":"publisher","DOI":"10.1002\/biot.201600193"},{"key":"e_1_2_10_15_1","doi-asserted-by":"publisher","DOI":"10.3390\/v12060662"},{"key":"e_1_2_10_16_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cobme.2021.100353"},{"key":"e_1_2_10_17_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtm.2021.02.010"},{"key":"e_1_2_10_18_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtm.2021.04.006"},{"key":"e_1_2_10_19_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.chroma.2021.462210"},{"key":"e_1_2_10_20_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtn.2021.06.026"},{"key":"e_1_2_10_21_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-021-21935-5"},{"key":"e_1_2_10_22_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jviromet.2013.10.038"},{"key":"e_1_2_10_23_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.langmuir.3c00241"},{"key":"e_1_2_10_24_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.omtm.2020.06.004"},{"key":"e_1_2_10_25_1","doi-asserted-by":"publisher","DOI":"10.1089\/biores.2020.0031"},{"key":"e_1_2_10_26_1","doi-asserted-by":"publisher","DOI":"10.2217\/fvl-2017-0011"},{"key":"e_1_2_10_27_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.neulet.2019.134310"},{"key":"e_1_2_10_28_1","doi-asserted-by":"publisher","DOI":"10.3390\/ijms20174133"},{"key":"e_1_2_10_29_1","doi-asserted-by":"publisher","DOI":"10.1080\/01926230701320337"},{"key":"e_1_2_10_30_1","doi-asserted-by":"publisher","DOI":"10.1128\/JVI.74.20.9441-9450.2000"}],"container-title":["Biotechnology Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/analyticalsciencejournals.onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/biot.202300667","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,8]],"date-time":"2025-10-08T19:26:36Z","timestamp":1759951596000},"score":1,"resource":{"primary":{"URL":"https:\/\/analyticalsciencejournals.onlinelibrary.wiley.com\/doi\/10.1002\/biot.202300667"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,3]]},"references-count":29,"journal-issue":{"issue":"3","published-print":{"date-parts":[[2024,3]]}},"alternative-id":["10.1002\/biot.202300667"],"URL":"https:\/\/doi.org\/10.1002\/biot.202300667","archive":["Portico"],"relation":{},"ISSN":["1860-6768","1860-7314"],"issn-type":[{"value":"1860-6768","type":"print"},{"value":"1860-7314","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,3]]},"article-number":"2300667"}}