1. Highly efficient gene tagging in the bryophyte Physcomitrella patens using the tobacco (Nicotiana tabacum) Tnt1 retrotransposon
- Author
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Julien Daniel, Fabien Nogué, Aline Epert, Corinne Mhiri, Josep M. Casacuberta, Daniel F. Voytas, Cristina Vives, Marie-Angèle Grandbastien, Florence Charlot, Beatriz Contreras, Ministerio de Ciencia e Innovación (España), National Institute for Basic Biology (Japan), Center for Research in Agricultural Genomics, Institut Jean-Pierre Bourgin (IJPB), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Department of Genetics - Cell Biology & Development and Center for Genome Engineering, University of Minnesota System, and Ministerio de Ciencia y Innovacion [BFU2009-11932, AGL2013-43244-R]
- Subjects
0106 biological sciences ,0301 basic medicine ,Retroelements ,Transcription, Genetic ,Physiology ,Nicotiana tabacum ,[SDV]Life Sciences [q-bio] ,Mutant ,Retrotransposon ,Plant Science ,Gene mutation ,Physcomitrella patens ,01 natural sciences ,03 medical and health sciences ,Transformation, Genetic ,Gene Expression Regulation, Plant ,Tobacco ,Tnt1 ,gene tagging ,Gene ,Selectable marker ,Genetics ,Polymorphism, Genetic ,biology ,Base Sequence ,fungi ,food and beverages ,biology.organism_classification ,Reverse genetics ,Bryopsida ,mutant population ,Mutagenesis, Insertional ,030104 developmental biology ,Genetic Techniques ,010606 plant biology & botany - Abstract
Because of its highly efficient homologous recombination, the moss Physcomitrella patens is a model organism particularly suited for reverse genetics, but this inherent characteristic limits forward genetic approaches. Here, we show that the tobacco (Nicotiana tabacum) retrotransposon Tnt1 efficiently transposes in P. patens, being the first retrotransposon from a vascular plant reported to transpose in a bryophyte. Tnt1 has a remarkable preference for insertion into genic regions, which makes it particularly suited for gene mutation. In order to stabilize Tnt1 insertions and make it easier to select for insertional mutants, we have developed a two-component system where a mini-Tnt1 with a retrotransposition selectable marker can only transpose when Tnt1 proteins are co-expressed from a separate expression unit. We present a new tool with which to produce insertional mutants in P. patens in a rapid and straightforward manner that complements the existing molecular and genetic toolkit for this model species., Work done at CRAG was supported by the Ministerio de Ciencia y Innovación (grants: BFU2009-11932 and AGL2013-43244-R). We thank Mitsuyasu Hasebe (National Institute for Basic Biology, Okazaki, Japan) for providing the p35S-loxP-Zeo vector.
- Published
- 2016
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