{"title":"Selection of a histidine auxotrophic <i>Marchantia polymorpha</i> strain with an auxotrophic selective marker.","authors":"Tatsushi Fukushima, Yutaka Kodama","doi":"10.5511/plantbiotechnology.22.0810a","DOIUrl":null,"url":null,"abstract":"<p><p><i>Marchantia polymorpha</i> has emerged as a model liverwort species, with molecular tools increasingly available. In the present study, we developed an auxotrophic strain of <i>M. polymorpha</i> and an auxotrophic selective marker gene as new experimental tools for this valuable model system. Using CRISPR (clustered regularly interspaced palindromic repeats)/Cas9-mediated genome editing, we mutated the genomic region for <i>IMIDAZOLEGLYCEROL-PHOSPHATE DEHYDRATASE</i> (<i>IGPD</i>) in <i>M. polymorpha</i> to disrupt the biosynthesis of histidine (<i>igpd</i>). We modified an <i>IGPD</i> gene (<i>IGPDm</i>) with silent mutations, generating a histidine auxotrophic selective marker gene that was not a target of our CRISPR/Cas9-mediated genome editing. The <i>M. polymorpha</i> <i>igpd</i> mutant was a histidine auxotrophic strain, growing only on medium containing histidine. The <i>igpd</i> mutant could be complemented by transformation with the <i>IGPDm</i> gene, indicating that this gene could be used as an auxotrophic selective marker. Using the <i>IGPDm</i> marker in the <i>igpd</i> mutant background, we produced transgenic lines without the need for antibiotic selection. The histidine auxotrophic strain <i>igpd</i> and auxotrophic selective marker <i>IGPDm</i> represent new molecular tools for <i>M. polymorpha</i> research.</p>","PeriodicalId":20411,"journal":{"name":"Plant Biotechnology","volume":null,"pages":null},"PeriodicalIF":1.4000,"publicationDate":"2022-12-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10240916/pdf/plantbiotechnology-39-4-22.0810a.pdf","citationCount":"1","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Plant Biotechnology","FirstCategoryId":"5","ListUrlMain":"https://doi.org/10.5511/plantbiotechnology.22.0810a","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
引用次数: 1
Abstract
Marchantia polymorpha has emerged as a model liverwort species, with molecular tools increasingly available. In the present study, we developed an auxotrophic strain of M. polymorpha and an auxotrophic selective marker gene as new experimental tools for this valuable model system. Using CRISPR (clustered regularly interspaced palindromic repeats)/Cas9-mediated genome editing, we mutated the genomic region for IMIDAZOLEGLYCEROL-PHOSPHATE DEHYDRATASE (IGPD) in M. polymorpha to disrupt the biosynthesis of histidine (igpd). We modified an IGPD gene (IGPDm) with silent mutations, generating a histidine auxotrophic selective marker gene that was not a target of our CRISPR/Cas9-mediated genome editing. The M. polymorphaigpd mutant was a histidine auxotrophic strain, growing only on medium containing histidine. The igpd mutant could be complemented by transformation with the IGPDm gene, indicating that this gene could be used as an auxotrophic selective marker. Using the IGPDm marker in the igpd mutant background, we produced transgenic lines without the need for antibiotic selection. The histidine auxotrophic strain igpd and auxotrophic selective marker IGPDm represent new molecular tools for M. polymorpha research.
期刊介绍:
Plant Biotechnology is an international, open-access, and online journal, published every three months by the Japanese Society for Plant Biotechnology. The journal, first published in 1984 as the predecessor journal, “Plant Tissue Culture Letters” and became its present form in 1997 when the society name was renamed to Japanese Society for Plant Cell and Molecular Biology, publishes findings in the areas from basic- to application research of plant biotechnology. The aim of Plant Biotechnology is to publish original and high-impact papers, in the most rapid turnaround time for reviewing, on the plant biotechnology including tissue culture, production of specialized metabolites, transgenic technology, and genome editing technology, and also on the related research fields including molecular biology, cell biology, genetics, plant breeding, plant physiology and biochemistry, metabolic engineering, synthetic biology, and bioinformatics.