Direct transformation of the liverwort Marchantia polymorpha L. by particle bombardment using immature thalli developing from spores

Abstract

The liverwort, Marchantia polymorpha L., belongs to a group of basal land plants and is an emerging model for plant biology. We established a procedure to prepare sporangia of M. polymorpha under laboratory conditions by promoting its transition to reproductive development by far-red light irradiation. Here we report an improved direct transformation system of M. polymorpha using immature thalli developing from spores. Hygromycin-resistant transformants were obtained on selective media by transformation with a plasmid carrying the hygromycin-phosphotransferase gene (hpt) conferring hygromycin resistance in 4 weeks. The aminoglycoside-3''-adenyltransferase gene (aadA) conferring spectinomycin resistance was also successfully used as an additional selectable marker for nuclear transformation of M. polymorpha. The availability of the aadA gene in addition to the hpt gene should make M. polymorpha a versatile host for genetic manipulation. DNA gel-blot analyses indicated that transformed thalli carried a variable number of copies of the transgene integrated into the genome. Although the previous system using thalli grown from gemmae required a two-step selection in liquid and solid media for 8 weeks, the system reported here using thalli developing from spores allows generation of transformants in half the time by direct selection on solid media, facilitating genetic analyses in this model plant.

Strigolactones regulate protonema branching and act as a quorum sensing-like signal in the moss Physcomitrella patens

“…These data and our demonstration that strigolactones are signalling molecules in the bryophyte P. patens support the hypothesis that strigolactones are very ancient molecules that have played a crucial role in the ecological adaptation of plants to terrestrial environment. Further studies with Marchantia, a mycorrhizal liverwort recently used in biological studies (Bowman et al, 2007;Chiyoda et al, 2008;Ishizaki et al, 2008,) would confirm the role of strigolactones in the establishment of arbuscular mycorrizal symbiosis in bryophytes.…”

Stable Transformation of Ferns Using Spores as Targets: Pteris vittata and Ceratopteris thalictroides

“…Stable genetic transformation has been accomplished in a few species outside angiosperms and gymnosperms, especially among bryophytes (Schaefer et al, 1991;Chiyoda et al, 2008;Ishizaki et al, 2008), but never in the Pteridophyta. Transient transformation methods have been developed both in Ceratopteris richardii 'C-fern' and P. vittata, which were limited to heterologous expression in gametophytes (Rutherford et al, 2004;Indriolo et al, 2010).…”

Co-option of a photoperiodic growth-phase transition system during land plant evolution

“…1). Exposure to far-red light (FR) is indispensable for this process 19 , suggesting the presence of light-quality-and day-length-dependent regulatory mechanisms in M. polymorpha. Interestingly, and in contrast to the aforementioned information, analysis of genome and transcriptome databases of M. polymorpha revealed single-copy orthologues for GI and FKF1, designated as MpGI and MpFKF, respectively (Fig.…”

“…Liverworts represent a critical point in the early evolution of land plants, especially the water-toland transition 17,18 . In addition, molecular genetic tools including transformation techniques [19][20][21][22] and strategies for targeted genome modification 23,24 have been established for M. polymorpha. Here, we show that the GI-FKF1 complex is present in the gametophyte generation of M. polymorpha, and that it plays an important role in regulating photoperiodic growth-phase transition in this generation.…”