biostudies-literatureUnknown5(4)Xiang HTobacco (<i>Nicotiana tabacum</i>) plants synthesize the psychoactive pyridine alkaloid nicotine, which has sparked growing interest in reducing nicotine levels through genome editing aiming at inactivating key biosynthetic genes. Although stable transformation-mediated genome editing is effective in tobacco, its polyploid nature complicates the complete knockout of genes and the segregation of transgenes from edited plants. In this study, we developed a non-transgenic genome editing method in tobacco by delivering the CRISPR/Cas machinery via an engineered negative-strand RNA rhabdovirus vector, followed by the regeneration of mutant plants through tissue culture. Using this method, we targeted six <i>berberine bridge enzyme-like protein</i> (<i>BBL</i>) family genes for mutagenesis, whicaBIOTECH449-464https://www.ebi.ac.uk/biostudies/studies/S-EPMC11624166biostudies-literatureDevelopment of an RNA virus vector for non-transgenic genome editing in tobacco and generation of &lt;i&gt;berberine bridge enzyme-like&lt;/i&gt; mutants with reduced nicotine content.PMC11624166Gao QJiang JLi ZWang SHuang HMi QXiang HNi SChen BZeng WKong WfalseDevelopment of an RNA virus vector for non-transgenic genome editing in tobacco and generation of &lt;i&gt;berberine bridge enzyme-like&lt;/i&gt; mutants with reduced nicotine content.Tobacco (<i>Nicotiana tabacum</i>) plants synthesize the psychoactive pyridine alkaloid nicotine, which has sparked growing interest in reducing nicotine levels through genome editing aiming at inactivating key biosynthetic genes. Although stable transformation-mediated genome editing is effective in tobacco, its polyploid nature complicates the complete knockout of genes and the segregation of transgenes from edited plants. In this study, we developed a non-transgenic genome editing method in tobacco by delivering the CRISPR/Cas machinery via an engineered negative-strand RNA rhabdovirus vector, followed by the regeneration of mutant plants through tissue culture. Using this method, we targeted six <i>berberine bridge enzyme-like protein</i> (<i>BBL</i>) family genes for mutagenesis, whic2024-01-01T00:00:00Z2024 Dec2025-04-04T23:31:36.119Z2025-04-04T23:31:36.119ZS-EPMC116241663965014210.1007/s42994-024-00188-y