Vovener de Verlands Edmond, Pamela A. Moon, Matthew Bremgartner, Xingbo Wu, Elias Bassil
{"title":"Agrobacterium-mediated transformation, selection and regeneration of Vanilla pompona","authors":"Vovener de Verlands Edmond, Pamela A. Moon, Matthew Bremgartner, Xingbo Wu, Elias Bassil","doi":"10.1007/s11240-024-02836-z","DOIUrl":null,"url":null,"abstract":"<p>Vanilla is a high-value tropical orchid cultivated for its aromatic fruit capsules that are used in foods, perfumes, and industrial products. <i>Vanilla planifolia</i> (Jacks ex. Andrews) is the most important commercially grown species, but its production is constrained by poor yield, variable quality, low genetic diversity and limited horticultural advancements. A closely related species, <i>Vanilla pompona</i> Schiede, characteristics which could be useful in breeding improved varieties: large fruit, potent aroma, and resistance to <i>Fusarium oxysporum</i> f. sp. <i>vanilla</i>. Here we describe tissue culture-based regeneration and <i>Agrobacterium</i>-mediated stable transformation systems for <i>V</i>. <i>pompona.</i> Vegetatively propagated tissue was used to test the efficacy of hygromycin and phosphinothricin selection and to assess the efficiency of three <i>Agrobacterium tumefaciens</i> strains (EHA105, AGL1.1, GV3101) in transformation protocols. Results revealed that for <i>V. pompona</i>, kanamycin is not an effective selection marker, whereas hygromycin and phosphinothricin can be used for screening transformants. AGL1.1 provided the highest transformation efficiency (37%) as compared to strains EHA105 (11%) and GV3101 (4%). Additionally, we incorporated the use of firefly luciferase as a visual reporter of transformation and were able to demonstrate that it provides more robust assessment than that of green fluorescent protein. Finally, we report a novel quantitative imaging method to assess the growth responses of <i>V</i>. <i>pompona</i> protocorm-like bodies in response to selection that could be useful to other plant transformation and selection efforts.</p>","PeriodicalId":20219,"journal":{"name":"Plant Cell, Tissue and Organ Culture","volume":"198 1","pages":""},"PeriodicalIF":2.3000,"publicationDate":"2024-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Plant Cell, Tissue and Organ Culture","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1007/s11240-024-02836-z","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Vanilla is a high-value tropical orchid cultivated for its aromatic fruit capsules that are used in foods, perfumes, and industrial products. Vanilla planifolia (Jacks ex. Andrews) is the most important commercially grown species, but its production is constrained by poor yield, variable quality, low genetic diversity and limited horticultural advancements. A closely related species, Vanilla pompona Schiede, characteristics which could be useful in breeding improved varieties: large fruit, potent aroma, and resistance to Fusarium oxysporum f. sp. vanilla. Here we describe tissue culture-based regeneration and Agrobacterium-mediated stable transformation systems for V. pompona. Vegetatively propagated tissue was used to test the efficacy of hygromycin and phosphinothricin selection and to assess the efficiency of three Agrobacterium tumefaciens strains (EHA105, AGL1.1, GV3101) in transformation protocols. Results revealed that for V. pompona, kanamycin is not an effective selection marker, whereas hygromycin and phosphinothricin can be used for screening transformants. AGL1.1 provided the highest transformation efficiency (37%) as compared to strains EHA105 (11%) and GV3101 (4%). Additionally, we incorporated the use of firefly luciferase as a visual reporter of transformation and were able to demonstrate that it provides more robust assessment than that of green fluorescent protein. Finally, we report a novel quantitative imaging method to assess the growth responses of V. pompona protocorm-like bodies in response to selection that could be useful to other plant transformation and selection efforts.
期刊介绍:
This journal highlights the myriad breakthrough technologies and discoveries in plant biology and biotechnology. Plant Cell, Tissue and Organ Culture (PCTOC: Journal of Plant Biotechnology) details high-throughput analysis of gene function and expression, gene silencing and overexpression analyses, RNAi, siRNA, and miRNA studies, and much more. It examines the transcriptional and/or translational events involved in gene regulation as well as those molecular controls involved in morphogenesis of plant cells and tissues.
The journal also covers practical and applied plant biotechnology, including regeneration, organogenesis and somatic embryogenesis, gene transfer, gene flow, secondary metabolites, metabolic engineering, and impact of transgene(s) dissemination into managed and unmanaged plant systems.