title: Conditional Gene Expression/Deletion Systems for Marchantia polymorpha Using its Own Heat-Shock Promoter and Cre/loxP-Mediated Site-Specific Recombination.
abstract: The liverwort Marchantia polymorpha is an emerging model plant suitable for addressing, using genetic approaches, various evolutionary questions in the land plant lineage. Haploid dominancy in its life cycle facilitates genetic analyses, but conversely limits the ability to isolate mutants of essential genes. To overcome this issue and to be employed in cell lineage, mosaic and cell autonomy analyses, we developed a system that allows conditional gene expression and deletion using a promoter of a heat-shock protein (HSP) gene and the Cre/loxP site-specific recombination system. Because the widely used promoter of the Arabidopsis HSP18.2 gene did not operate in M. polymorpha, we identified a promoter of an endogenous HSP gene, MpHSP17.8A1, which exhibited a highly inducible transient expression level upon heat shock with a low basal activity level. Reporter genes fused to this promoter were induced globally in thalli under whole-plant heat treatment and also locally using a laser-assisted targeted heating technique. By expressing Cre fused to the glucocorticoid receptor under the control of the MpHSP17.8A1 promoter, a low background, sufficiently inducible control for loxP-mediated recombination could be achieved in M. polymorpha. Based on these findings, we developed a Gateway technology-based binary vector for the conditional induction of gene deletions.
authors.affiliation: Graduate School of Biostudies, Kyoto University, Kyoto, 606-8502 Japan.
authors.forename: Ryuichi
authors.initials: R
authors.lastname: Nishihama
authors.affiliation: Graduate School of Biostudies, Kyoto University, Kyoto, 606-8502 Japan.
authors.forename: Sakiko
authors.initials: S
authors.lastname: Ishida
authors.affiliation: Faculty of Education, Gifu Shotoku Gakuen University, Gifu, 501-6194 Japan.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Urawa
authors.affiliation: Spectrography and Bioimaging Facility, NIBB Core Facilities, National Institute for Basic Biology, Okazaki, Aichi, 444-8585 Japan Department of Basic Biology in the School of Life Science, SOKENDAI (the Graduate University for Advanced Studies), Okazaki, Aichi, 444-8585 Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kamei
authors.affiliation: Graduate School of Biostudies, Kyoto University, Kyoto, 606-8502 Japan tkohchi@lif.kyoto-u.ac.jp.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Kohchi
mesh_terms: D017353:Gene Deletion; D018506:Gene Expression Regulation, Plant; D018390:Gene Targeting; D005822:Genetic Vectors / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism; D018869:Heat-Shock Response; D019426:Integrases / Q000378:metabolism*; D007834:Lasers; D047008:Marchantia / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011995:Recombination, Genetic
chemical_list: D006360:Heat-Shock Proteins; C045073:Cre recombinase; D019426:Integrases
keywords: Conditional approach; Cre/loxP site-specific recombination; Glucocorticoid receptor; Heat-shock protein; Liverwort; Marchantia polymorpha