Transient and stable transformation of Ceratopteris richardii gametophytes

BMC Research Notes

Chi-Lien Cheng¹  Erin E Irish¹  Angela R Cordle¹  Linh T Bui² 

[1] Department of Biology, University of Iowa, Iowa City 52242, IA, USA;Donald Danforth Plant Science Center, St. Louis 63132, MO, USA

关键词: Hygromycin;    GUS;    GFP;    Cellulose;    Regeneration;    Agrobacterium;    Gametophyte;    Fern;    Ceratopteris;   

Others  :  1232441
DOI  :  10.1186/s13104-015-1193-x

 received in 2014-12-18, accepted in 2015-05-20,  发布年份 2015

【 摘 要 】

Background

Ferns, being vascular yet seedless, present unparalleled opportunities to investigate important questions regarding the evolution and development of land plants. Ceratopteris richardii, a diploid, homosporous fern has been advanced as a model fern system; however, the tenuous ability to transform the genome of this fern greatly limited its usefulness as a model organism. Here we report a simple and reliable Agrobacterium-mediated method for generating transient and stable transformants of mature C. richardii gametophytes.

Results

Transformation success was achieved by enzyme treatment that partially digested the cell walls of mature gametophytes to facilitate Agrobacteria infection. Co-incubation of Agrobacteria with enzymatically treated gametophytes was sufficient to generate transient transformants at a frequency of nearly 90% under optimal conditions. Stable transformation was achieved at a rate of nearly 3% by regenerating entire gametophytes from single transformed cells from T₀ gametophytes on selective media.

Conclusions

This transformation method will allow for the immediate observation of phenotypes in the haploid gametophytes of transformed plants, as well as the generation of stably transformed C. richardii lines for further analysis. Transformation capability will greatly facilitate gene functional studies in C. richardii, more fully realizing the potential of this model fern species. These protocols may be adapted to other plant species that are recalcitrant to Agrobacterium-mediated transformation.

【 授权许可】

   
2015 Bui et al.

【 图 表 】

Figure1.

Figure2.

Figure3.

Figure4.

【 参考文献 】

• [1]Fernández H, Revilla M: In vitro culture of ornamental ferns. Plant Cell Tissue Organ Cult 2003, 73(1):1-13.
• [2]Hickok LG, Warne TR, Slocum MK: Ceratopteris richardii—application for experimental plant biology. Am J Bot 1987, 74(8):1304-1316.
• [3]Dyer AF (1979) The experimental biology of ferns. Academic, London
• [4]Sheffield E, Bell P: Ultrastructural aspects of sporogenesis in a fern, Pteridium aquilinum (L.) Kuhn. Ann Bot 1979, 44(4):393-405.
• [5]Cordle AR, Bui LT, Irish EE, Cheng C-L: Laboratory-induced apogamy and apospory in Ceratopteris richardii. In Working with ferns. Edited by Fernadez H, Kumar A, Revilla MA. Springer, New York; 2010:25-36.
• [6]Banks JA: Gametophyte development in ferns. Annu Rev Plant Biol 1999, 50(1):163-186.
• [7]Hickok LG, Warne TR, Fribourg RS: The biology of the fern Ceratoperis and its use as a model system. Int J Dev Biol 1995, 156(3):332-345.
• [8]Banks JA: Sex determination in the fern Ceratopteris. Trends Plant Sci 1997, 2(5):175-180.
• [9]Edwards ES, Roux SJ: Gravity and light control of the developmental polarity of regenerating protoplasts isolated from prothallial cells of the fern Ceratopteris richardii. Plant Cell Rep 1998, 17(9):711-716.
• [10]Salmi ML, Roux SJ: Gene expression changes induced by space flight in single-cells of the fern Ceratopteris richardii. Planta 2008, 229(1):151-159.
• [11]DeYoung B, Weber T, Hass B, Banks JA: Generating autotetraploid sporophytes and their use in analyzing mutations affecting gametophyte development in the fern Ceratopteris. Genetics 1997, 147(2):809-814.
• [12]Cordle AR, Irish EE, Cheng CL: Apogamy induction in Ceratopteris richardii. Int J Plant Sci 2007, 168(4):361-369.
• [13]Bui LT, Hurst A, Irish EE, Cheng CL: The effects of sugars and ethylene on apospory and regeneration in Ceratopteris richardii. Am J Plant Sci 2012, 3(7):953-961.
• [14]Leroux O, Eeckhout S, Viane RL, Popper ZA: Ceratopteris richardii (C-fern): a model for investigating adaptive modification of vascular plant cell walls. Front Plant Sci 2013, 4:367.
• [15]Rutherford G, Tanurdzic M, Hasebe M, Banks JA: A systemic gene silencing method suitable for high throughput, reverse genetic analyses of gene function in fern gametophytes. BMC Plant Biol 2004, 4(1):6. BioMed Central Full Text
• [16]Indriolo E, Na G, Ellis D, Salt DE, Banks JA: A vacuolar arsenite transporter necessary for arsenic tolerance in the arsenic hyperaccumulating fern Pteris vittata is missing in flowering plants. Plant Cell 2010, 22(6):2045-2057.
• [17]Stout SC, Clark GB, Archer-Evans S, Roux SJ: Rapid and efficient suppression of gene expression in a single-cell model system, Ceratopteris richardii. Plant Physiol 2003, 131(3):1165-1168.
• [18]Stout SC (2004) Physiological and bioinformatics studies on polarity development in Ceratopteris richardii spores. Ph.D. thesis. The University of Texas at Austin
• [19]Muthukumar B, Joyce BL, Elless MP, Stewart CN: Stable transformation of ferns using spores as targets: Pteris vittata and Ceratopteris thalictroides. Plant Physiol 2013, 163(2):648-658.
• [20]Plackett A, Huang L, Sanders HL, Langdale JA: High-efficiency stable transformation of the model fern species Ceratopteris richardii via microparticle bombardment. Plant Physiol 2014, 165:3-14.
• [21]Banks JA: Sex-determining genes in the homosporous fern Ceratopteris. Development 1994, 120(7):1949-1958.
• [22]Curtis MD, Grossniklaus U: A gateway cloning vector set for high-throughput functional analysis of genes in Planta. Plant Physiol 2003, 133(2):462-469.
• [23]Bekkaoui F, Datla RSS, Pilon M, Tautorus TE, Crosby WL, Dunstan DI: The effects of promoter on transient expression in conifer cell lines. Theor Appl Genet 1990, 79(3):353-359.
• [24]Ganger MT, Girouard JA, Smith HM, Bahny BA, Ewing SJ: Antheridiogen and abscisic acid affect conversion and ANI1 expression in Ceratopteris richardii gametophytes. Botany 2014, 93(2):109-116.
• [25]Kohli A, Twyman RM, Abranches R, Wegel E, Stoger E, Christou P: Transgene integration, organization and interaction in plants. Plant Mol Biol 2003, 52(2):247-258.
• [26]Preacher KJ (2001) Calculation for the Chi square test: an interactive calculation tool for Chi square tests of goodness of fit and independence [Computer software]. Available from http://quantpsy.org
• [27]Utermark J, Karlovsky P (2008) Genetic transformation of filamentous fungi by Agrobacterium tumefaciens. Nat Protoc 83
• [28]Jefferson RA, Kavanagh TA, Bevan MW: GUS fusions: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 1987, 6(13):3901.
• [29]Ishizaki K, Chiyoda S, Yamato KT, Kohchi T: Agrobacterium-mediated transformation of the haploid liverwort Marchantia polymorpha L., an emerging model for plant biology. Plant Cell Physiol 2008, 49(7):1084-1091.
• [30]Kiefer E, Heller W, Ernst D: A simple and efficient protocol for isolation of functional RNA from plant tissues rich in secondary metabolites. Plant Mol Biol Rep 2000, 18(1):33-39.