title: A Cyan Fluorescent Reporter Expressed from the Chloroplast Genome of Marchantia polymorpha.
abstract: Recently, the liverwort Marchantia polymorpha has received increasing attention as a basal plant model for multicellular studies. Its ease of handling, well-characterized plastome and proven protocols for biolistic plastid transformation qualify M. polymorpha as an attractive platform to study the evolution of chloroplasts during the transition from water to land. In addition, chloroplasts of M. polymorpha provide a convenient test-bed for the characterization of genetic elements involved in plastid gene expression due to the absence of mechanisms for RNA editing. While reporter genes have proven valuable to the qualitative and quantitative study of gene expression in chloroplasts, expression of green fluorescent protein (GFP) in chloroplasts of M. polymorpha has proven problematic. We report the design of a codon-optimized gfp varian, mturq2cp, which allowed successful expression of a cyan fluorescent protein under control of the tobacco psbA promoter from the chloroplast genome of M. polymorpha. We demonstrate the utility of mturq2cp in (i) early screening for transplastomic events following biolistic transformation of M. polymorpha spores; (ii) visualization of stromules as elements of plastid structure in Marchantia; and (iii) quantitative microscopy for the analysis of promoter activity.
authors.affiliation: Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK.
authors.forename: Christian R
authors.initials: CR
authors.lastname: Boehm
authors.affiliation: Department of Botany, Graduate School of Science, Kyoto University, Sakyo-ku, Kyoto, 606-8502 Japan Present address: RIKEN Center for Sustainable Resource Science, 1-7-22 Suehiro, Tsurumi, Yokohama, 230-0045 Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Ueda
authors.affiliation: Department of Botany, Graduate School of Science, Kyoto University, Sakyo-ku, Kyoto, 606-8502 Japan.
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Nishimura
authors.affiliation: Department of Botany, Graduate School of Science, Kyoto University, Sakyo-ku, Kyoto, 606-8502 Japan CREST, Japan Science and Technology Agency, Chiyoda-ku, Tokyo, 102-0076 Japan.
authors.forename: Toshiharu
authors.initials: T
authors.lastname: Shikanai
authors.affiliation: Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK jh295@cam.ac.uk.
authors.forename: Jim
authors.initials: J
authors.lastname: Haseloff
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D005453:Fluorescence; D054628:Genome, Chloroplast; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D047008:Marchantia / Q000235:genetics*; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified; D014170:Transformation, Genetic
chemical_list: C485182:Cyan Fluorescent Protein; D049452:Green Fluorescent Proteins
keywords: Chloroplast; Codon bias; GFP; Marchantia polymorpha; Reporter gene; Stromules