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Using TAQing2.0 and conventional mutagenesis (by nitrosoguanidine), we obtained mutant asexual <jats:styled-content style=\"fixed-case\"><jats:italic>Candida utilis<\/jats:italic><\/jats:styled-content> strains capable of growing under highly acidic conditions (pH\u20091.8). Whole genome resequencing revealed that the genomic sequences of mutants generated by both methods contain a negligible small population of unmappable sequences, suggesting that both types of mutants can be regarded as equivalent to naturally occurring mutants. TAQing2.0 mutants exhibit multiple genome rearrangements with few point mutations, whereas conventional mutagenesis produces numerous point mutations. This feature enabled us to easily identify candidate genes (e.g., <jats:italic>LYP<\/jats:italic><jats:italic>1<\/jats:italic> homolog) responsible for acid resistance. TAQing2.0 is a powerful and versatile tool for mutant production and gene hunting without invasion of foreign DNA.<\/jats:p>","DOI":"10.1111\/gtc.70010","type":"journal-article","created":{"date-parts":[[2025,3,11]],"date-time":"2025-03-11T23:19:52Z","timestamp":1741735192000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Advantages of Mutant Generation by Genome Rearrangements of Non\u2010Conventional Yeast via Direct Nuclease Transfection"],"prefix":"10.1111","volume":"30","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3797-4410","authenticated-orcid":false,"given":"Arisa H.","family":"Oda","sequence":"first","affiliation":[{"name":"Department of Life Sciences Graduate School of Arts &amp; Sciences, the University of Tokyo  Tokyo Japan"},{"name":"Collaborative Research Institute for Innovative Microbiology  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Taishi","family":"Yasukawa","sequence":"additional","affiliation":[{"name":"Mitsubishi Corporation Life Sciences Limited  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Miki","family":"Tamura","sequence":"additional","affiliation":[{"name":"Department of Life Sciences Graduate School of Arts &amp; Sciences, the University of Tokyo  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ayumu","family":"Sano","sequence":"additional","affiliation":[{"name":"Mitsubishi Corporation Life Sciences Limited  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Naohisa","family":"Masuo","sequence":"additional","affiliation":[{"name":"Mitsubishi Corporation Life Sciences Limited  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kunihiro","family":"Ohta","sequence":"additional","affiliation":[{"name":"Department of Life Sciences Graduate School of Arts &amp; Sciences, the University of Tokyo  Tokyo Japan"},{"name":"Collaborative Research Institute for Innovative Microbiology  Tokyo Japan"},{"name":"The Universal Biology Institute of the University of Tokyo  Tokyo Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2025,3,11]]},"reference":[{"key":"e_1_2_9_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/0006\u2010291X(65)90855\u20107"},{"key":"e_1_2_9_3_1","doi-asserted-by":"publisher","DOI":"10.1101\/pdb.top082651"},{"key":"e_1_2_9_4_1","doi-asserted-by":"publisher","DOI":"10.1038\/s42003\u2010023\u201005285\u20100"},{"issue":"3","key":"e_1_2_9_5_1","first-page":"407","article-title":"Production of Food Grade Yeasts","volume":"44","author":"Bekatorou A.","year":"2006","journal-title":"Food Technology and Biotechnology"},{"key":"e_1_2_9_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/0027\u20105107(70)90148\u2010X"},{"key":"e_1_2_9_7_1","doi-asserted-by":"publisher","DOI":"10.1186\/1471\u20102105\u201010\u2010421"},{"key":"e_1_2_9_8_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.ygeno.2020.12.004"},{"key":"e_1_2_9_9_1","doi-asserted-by":"publisher","DOI":"10.1038\/nmeth.4035"},{"key":"e_1_2_9_10_1","doi-asserted-by":"publisher","DOI":"10.1002\/0471142727.mb1312s23"},{"key":"e_1_2_9_11_1","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2007.13"},{"key":"e_1_2_9_12_1","first-page":"87","volume-title":"Methods in Enzymology","author":"Gietz R. 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