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

Muthukumar, B.; Joyce, Blake L.; Elless, Mark P.; Stewart, C. Neal

doi:10.1104/pp.113.224675

Cited by 46 publications

(42 citation statements)

References 57 publications

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“…1f) as in the peripheral zone of seed plants (Frank et al, 2015). There have been several recent advances in knockout technologies in ferns (Muthukumar et al, 2013;Plackett et al, 2014;Bui et al, 2015) and transcriptome analyses (Frank et al, 2015). The firm recognition of the fern and lycophyte meristems as multicellular with cytohistochemical zonation is integral for the interpretation of broad comparative developmental analyses.…”

Section: Resultsmentioning

confidence: 99%

Bringing the multicellular fern meristem into focus

Ambrose

Vasco

2016

New Phytologist

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Section: Resultsmentioning

confidence: 99%

Bringing the multicellular fern meristem into focus

Ambrose

Vasco

2016

New Phytologist

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“…By contrast, hygromycin selection was found to be a very efficient screening mechanism when identifying transformants regenerating from callus tissue. Regardless of the selection technique, A. tumefaciens-mediated transformation of both C. thalictroides and P. vittata spores occurred at very low frequencies (0.03% and 0.053%, respectively; Muthukumar et al, 2013). Attempts in the same report to directly transform a population of P. vittata spores by microparticle bombardment similarly yielded a very low transformation efficiency of only 0.012%.…”

Section: Discussionmentioning

confidence: 97%

“…Although past attempts to transform C. richardii via A. tumefaciens reportedly failed (Hickok et al, 1987), a protocol for successful A. tumefaciens-mediated transformation of C. thalictroides has recently been published (Muthukumar et al, 2013), with transformation achieved through infection of geminating spores. Interestingly, Muthukumar et al (2013) report that their attempts to use hygromycin as a selectable marker for transformation were unsuccessful. This is potentially explained through our observation that hygromycin killed untransformed T1 gametophytes on germination (Supplemental Fig.…”

Section: Discussionmentioning

confidence: 99%

“…The available data thus suggest that transformation of spores is an inherently less efficient method than transformation of callus tissue. Using the protocol described in this paper, the time from callus transformation to recovery of T1 spores was 18 and 16 weeks for C. richardii and C. thalictroides, respectively, in contrast to an 11-to 13-week period for C. thalictroides via A. tumefaciens-mediated transformation (Muthukumar et al, 2013). Despite the longer time frame, transformation of the sporophyte generation confers the advantage of direct production of T1 spores without an intervening recombination/outcrossing event (sexual reproduction of T0 gametophytes).…”

Section: Discussionmentioning

confidence: 99%

“…Transmission to subsequent generations was not determined. A recently published protocol utilizing Agrobacterium tumefaciens-mediated transformation of spores reported stable transformation in two fern species, P. vittata and Ceratopteris thalictroides (Muthukumar et al, 2013), but the very low transformation efficiencies achieved (0.053% and 0.03%, respectively) preclude routine adoption of this approach.…”

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confidence: 99%

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High-Efficiency Stable Transformation of the Model Fern Species Ceratopteris richardii via Microparticle Bombardment

et al. 2014

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Ferns represent the most closely related extant lineage to seed plants. The aquatic fern Ceratopteris richardii has been subject to research for a considerable period of time, but analyses of the genetic programs underpinning developmental processes have been hampered by a large genome size, a lack of available mutants, and an inability to create stable transgenic lines. In this paper, we report a protocol for efficient stable genetic transformation of C. richardii and a closely related species Ceratopteris thalictroides using microparticle bombardment. Indeterminate callus was generated and maintained from the sporophytes of both species using cytokinin treatment. In proof-of-principle experiments, a 35S::b-glucuronidase (GUS) expression cassette was introduced into callus cells via tungsten microparticles, and stable transformants were selected via a linked hygromycin B resistance marker. The presence of the transgene in regenerated plants and in subsequent generations was validated using DNA-blot analysis, reverse transcription-polymerase chain reaction, and GUS staining. GUS staining patterns in most vegetative tissues corresponded with constitutive gene expression. The protocol described in this paper yields transformation efficiencies far greater than those previously published and represents a significant step toward the establishment of a tractable fern genetic model.

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Plant Nanobionic Sensors for Arsenic Detection

et al. 2020

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Arsenic is a highly toxic heavy‐metal pollutant which poses a significant health risk to humans and other ecosystems. In this work, the natural ability of wild‐type plants to pre‐concentrate and extract arsenic from the belowground environment is exploited to engineer plant nanobionic sensors for real‐time arsenic detection. Near‐infrared fluorescent nanosensors are specifically designed for sensitive and selective detection of arsenite. These optical nanosensors are embedded in plant tissues to non‐destructively access and monitor the internal dynamics of arsenic taken up by the plants via the roots. The integration of optical nanosensors with living plants enables the conversion of plants into self‐powered autosamplers of arsenic from their environment. Arsenite detection is demonstrated with three different plant species as nanobionic sensors. Based on an experimentally validated kinetic model, the nanobionic sensor could detect 0.6 and 0.2 ppb levels of arsenic after 7 and 14 days respectively by exploiting the natural ability of Pteris cretica ferns to hyperaccumulate and tolerate exceptionally high level of arsenic. The sensor readout could also be interfaced with portable electronics at a standoff distance, potentially enabling applications in environmental monitoring and agronomic research.

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

Cited by 46 publications

References 57 publications

Bringing the multicellular fern meristem into focus

Bringing the multicellular fern meristem into focus

High-Efficiency Stable Transformation of the Model Fern Species Ceratopteris richardii via Microparticle Bombardment

Plant Nanobionic Sensors for Arsenic Detection

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