Advertisement

Plant Cell, Tissue and Organ Culture

, Volume 40, Issue 1, pp 97–100 | Cite as

Assessment of microinjection for introducing DNA into uninuclear microspores of rapeseed

  • Elizabeth Jones-Villeneuve
  • Bin Huang
  • Isabelle Prudhomme
  • Sharon Bird
  • Roger Kemble
  • Jiro Hattori
  • Brian Miki
Research Notes

Abstract

Approximately 2,000 embryogenic uninuclear microspores of rapeseed (Brassica napus) cv. Topas were intranuclearly injected with a chimaeric β-glucuronidase (Escherichia coli Uid A) gene. Stable integration had not occurred among 55 plants that were regenerated. Coinjection of the dye Lucifer Yellow and detection of injected DNA by the polymerase chain reaction revealed high frequencies of transfer. However, the amount of DNA injected was less than 20 copies, which may have been insufficient for stable transformation of microspores.

Key words

Brassica napus β-glucuronidase polymerase chain reaction transformation 

Abbreviations

PCR

polymerase chain reaction

GUS

β-glucuronidase

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Bolik M & Koop HU (1991) Identification of embryogenic microspores of barley (Hordeum vulgare L.) by individual selection and culture and their potential for transformation by microin-jection. Protoplasma 162: 61–68Google Scholar
  2. Crossway A, Oakes JV, Irvine JM, Ward B, Knauf VC & Shewmaker CK (1986) Integration of foreign DNA following microinjection of tobacco mesophyll protoplasts. Mol. Gen. Genet. 202: 179–185Google Scholar
  3. Gaillard A, Matthys-Rochon E & Dumas C (1992) Selection of microspore derived embryogenic structures in maize related to transformation potential by microinjection. Bot. Acta 105: 313–318Google Scholar
  4. Gilliland G, Perrin S & Franklin Bunn H (1990) Competitive PCR for quantitation of mRNA. In: Innis MA, Gelfand DH, Sninsky JJ & White TJ (Eds) PCR Protocols: A Guide to Methods and Applications (pp 60–69) Academic Press, TorontoGoogle Scholar
  5. Huang B & Keller WA (1989) Microspore culture technology. J. Tissue Cult. Meth. 12: 171–178Google Scholar
  6. Jefferson RA, Kavanaugh TA & Bevan MW (1987) GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in high plants. EMBO J. 6: 3901–3907Google Scholar
  7. Miki BL, Huang B, Bird S, Kemble R, Simmonds D & Keller W (1989) A procedure for the microinjection of plant cells and protoplasts. J. Tissue Cult. Meth. 12: 139–144Google Scholar
  8. Neuhaus G, Spangenberg G, Mittelsten Scheid O & Schwieger H-G (1987) Transgenic rapeseed plants obtained by the microinjection of DNA into microspore-derived embryos. Theor. Appl. Genet. 75: 30–36Google Scholar
  9. Nomura K & Komamine A (1986) Embryogenesis from microin-jected single cells in a carrot cell suspension culture. Plant Sci. 44: 53–58Google Scholar
  10. Reich TJ, Iyer VN & Miki BL (1986a) Efficient transformation of alfalfa protoplasts by the intranuclear microinjection of Ti plasmids. Bio/Technol 4: 1001–1004Google Scholar
  11. Reich T, Iyer VN, Scobie B & Miki BL (1986b) A detailed procedure for the intranuclear microinjection of plant protoplasts. Can J. Bot. 64: 1255–1258Google Scholar

Copyright information

© Kluwer Academic Publishers 1995

Authors and Affiliations

  • Elizabeth Jones-Villeneuve
    • 1
  • Bin Huang
    • 1
  • Isabelle Prudhomme
    • 1
  • Sharon Bird
    • 1
  • Roger Kemble
    • 1
  • Jiro Hattori
    • 2
  • Brian Miki
    • 2
  1. 1.Canola Research StationPioneer Hi-Bred Production LimitedGeorgetownCanada
  2. 2.Plant Research CentreAgriculture CanadaOttawaCanada

Personalised recommendations