Abstract
A procedure for culturing protoplasts from slowly growing embryogenic calli of wheat was developed. The procedure was dependent on the ability to isolate large numbers of culturable protoplasts from slowly growing embryogenic callus. Approximately 68% of the isolated protoplasts divided, and 22% formed colonies; of the latter, 67% continued to proliferate. Plating efficiency was reduced when protoplasts were transformed by polythylene glycol, electroporation, and/or Agrobacterium. Intact cells were also directly transformed by electroporation. Direct electroporation of the Agrobacterium binary vector into intact cells resulted in a significant increase of GUS activity over the control.
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Chibbar RN, Karatha KK, Leung NJ, Qureshi J, Caswell K (1991) Transient expression of marker genes in immature zygotic embryos of spring wheat (Triticum aestivum) through microprojectile bombardment. Genome 34:453–460
Gordon-Kamm WJ, Spencer TM, Mangano ML, Adams TR, Daines RJ, Start WG, O'Brien JV, Chambers SA, Adams WR, Willets NG, Rice TB, Mackey CJ, Kruger RW, Kausch AP, Lemaux PG (1990) Transformation of maize cells and regeneration of fertile transgenic plants. Plant Cell 2:603–618
Harris R, Wright M, Byrne M, Varnum J, Brightwell B, Schubert K (1989) Callus formation and plant regeneration from protoplasts derived from suspension cultures of wheat (Triticum aestivum L.). Plant Cell Rep 7:337–340
Hayashimoto A, Li Z, Murai N (1990) A polyethylene glycolmediated protoplast transformation system for production of fertile transgenic rice plants. Plant Physiol 93:857–863
Kao KN, Michayluk MR (1975) Nutritional requirements for growth of Vicia hajastana cells and protoplasts at a very low population density in liquid media. Planta 126:105–110
Klein TM, Kornstein L, Stanford JC, Fromm ME (1989) Genetic transformation of maize cells by particle bombardment. Plant Physiol 91:440–444
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–493
Potrykus I (1991) Gene transfer to plants: assessment of published approaches and results. Annu Rev Plant Physiol 42:205–225
Potrykus I, Saul MW, Petruska J, Paszkoski J, Shillito RD (1985) Direct gene transfer to cells of a graminaceous monocot. Mol Gen Genet 199:183–188
Rainieri DM, Bottino P, Gordon MP, Nester EW (1990) Agrobacterium-mediated transformation of rice Oryza sativa L. Bio/Technology 8:33–38
Rhodes CA, Pierce DA, Mettler U, Mascarenhas D, Detmer J (1988) Genetically transformed maize plants from protoplasts. Science 240:204–207
Shimamoto K, Terada R, Izawa T, Fujimoto H (1989) Fertile transgenic rice plants regenerated from transformed protoplasts. Nature 338:736–740
Skuzeski JM, Nicholes LM, Gestland RF (1990) Analysis of leaky viral translation termination codons in vitro by transient expression of improved ß-glucuronidase. Plant Mol Biol 15:65–79
Vasil V, Redway F, Vasil IK (1990) Regeneration of plants from embryogenic suspension culture protoplasts of wheat (Triticum aestivum L.). Bio/Technology 8:429–433
Vasil V, Brown SM, Re D, Fromm ME, Vasil IK (1991) Stably transformed callus lines from microprojectile bombardment of cell suspension cultures of wheat. Bio/Technology 10:667–674
Vasil V, Casillo AM, Fromm ME, Vasil IK (1992) Herbicide-resistant fertile wheat plants obtained by microprojectile bombardment of regenerable embryogenic callus. Bio/Technology 9:743–747
Zaghmount OMF, Torello WA (1990) Isolation and culture of protoplasts from embryogenic suspension cultures of red fescue (Festuca rubra L.). Plant Cell Rep 9:340–343
Zaghmout OMF, Trolinder NL (1993a) Factors affecting transient gene expression in protoplasts isolated from slowly growing embryogenic wheat callus. Theor Appl Genet 86:721–730
Zaghmout OMF, Trolinder NL (1993b) Simple and efficient method for directly electroporating Agrobacterium plasmid DNA into wheat callus cells. Nucleic Acids Res 21:1048
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Communicated by K. Tsunewaki
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Zaghmout, O.M.F. Transformation of protoplasts and intact cells from slowly growing embryogenic callus of wheat (Triticum aestivum L.). Theoret. Appl. Genetics 89, 577–582 (1994). https://doi.org/10.1007/BF00222451
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DOI: https://doi.org/10.1007/BF00222451