Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Transformation and regeneration of Brassica rapa using Agrobacterium tumefaciens


Transformation and regeneration procedures for obtaining transgenic Brassica rapa ssp. oleifera plants are described. Regeneration frequencies were increasedby using silver nitrate and by adjusting the duration of exposure to 2,4-D. For transformation, Agrobacterium tumefaciens strain EHA101 containing a binary plasmid with the neomycin phosphotransferase gene (NPT II) and the b-glucuronidase gene (GUS) was cocultivated with hypocotyl explants from the oilseed B. rapa cvs. Tobin and Emma. Transformed plants were obtained within three months of cocultivation. Transformation frequencies for the cultivars Tobin and Emma were 1–9%. Evidence for transformation was shown by NPT II dot blot assay, the GUS fluorometric assay, Southern analysis, and segregation of the kanamycin-resistance trait in the progeny. The transformation and regeneration procedure described here has been used routinely to transform two cultivars of B. rapa and 18 cultivars of B. napus.

This is a preview of subscription content, log in to check access.


  1. Altenbach SB, Kuo C-C, Staraci LC, Pearson KW, Wainwright C, Georgescu A, Townsend J (1992) Accumulation of a Brazil nut albumin in seeds of transgenic canola results in enhanced levels of seed protein methionine. Plant Mol Biol 18:235–246

  2. Barfield DG, Pua E-C (1991) Gene transfer in plants of Brassica juncea using Agrobacterium tumefaciens-mediated transformation. Plant Cell Rep 10:308–314

  3. Chi G-L, Barfield DG, Sim G-E, Pua E-C (1990) Effect of AgNO3 and aminoethoxyvinylglycine on in vitro shoot and root organogenesis from seedling explants of recalcitrant Brassica genotypes. Plant Cell Rep 9:195–198

  4. Chi G-L, Pua E-C (1989) Ethylene inhibitors enhanced de novo shoot regeneration from cotyledons of Brassica campestris ssp. chinensis (Chinese cabbage) in vitro. Plant Sci 64:243–250

  5. Comai L, Moran P, Maslyar D (1990) Novel and useful properties of a chimeric plant promoter combining CaMV 35S and MAS elements. Plant Molec Biol 15:373–381

  6. De Block M, De Brouwer D, Terming P (1989) Transformation of Brassicanapus and Brassica oleracea using Agrobacterium tumefaciens and the expression of the bar and neo genes in the transgenic plants. Plant Physiol 91:694–701

  7. Dellaporta SL, Wood J, Hicks JB (1983) A plant DNA minipreparation; version II. Plant Mol Biol Rep 1:19–21

  8. Facciotti D, Turner J, Justicia R, Radke S (1989) Somatic regeneration and recovery of trans genic plants from Brassica campestris. In: Quiros CF and McGuire PE (eds) Proceedings of the Fifth Cracifer Genetics Workshop, Univ Calif Genet. Res Conserv Prog Oakland, CA (Rep No 4) p. 48

  9. Fry J, Barnason A, Horsch RB (1987) Transformation of Brassica napus with Agrobacterium tumefaciens based vectors. Plant Cell Rep 6:321–325

  10. Hachey JE, Sharma KK, Moloney MM (1991) Efficient shoot regeneration of Brassica campestris using cotyledon explants cultured in vitro. Plant Cell Rep 9:549–554

  11. Hood EE, Helmer GL, Fraley RT, Chilton M-D (1986) The hypervirulence of Agrobacterium tumefaciens A281 is encoded in a region of pTiBo542 outside of T-DNA. J Bacteriol 168:1291–1301

  12. Jefferson RA (1987) Assaying chimeric genes in plants: the GUS gene fusion system. Plant Mol Biol Rep 5:387–405

  13. Jorgensen R, Snyder C, Jones JDG (1987) T-DNA is organized predomi-nantly in inverted repeat structures in plants transformed with Agrobacterium tumefaciens C58 derivitives. Mol Gen Genet 207:471–477

  14. Knutzon DS, Thompson GA, Radke SE, Johnson WB, Knauf VC, Kridl JC (1992) Modification of Brassica seed oil by antisense expression of a stearoyl-acyl carrier protein desaturase gene. Proc Natl Acad Sci USA 89:2624–2628

  15. Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York

  16. McBride KE, Summerfelt KR (1990) Improved vectors for Agrobacterium-mediated plant transformation. Plant Mol Biol 14:269–276

  17. Murata M, Orton TJ (1987) Callus initiation and regeneration capacities in Brassica species. Plant Cell, Tiss and Organ Cult 11:111–123

  18. Nakanishi T, Hinata K (1973) An effective time for CO2 gas treatment in overcoming self-incompatibility in Brassica. Plant Cell Physiol 14:873–879

  19. Pua E-C, Mehra-Palta A, Nagy F, Chua N-H (1987) Transgenic plants of Brassica napus L. Bio/Technology 5:815–817

  20. Purnhauser L, Medgyesy P, Czako M, Dix PJ, Marton L (1987) Stimulation of shoot regeneration in Triticum aestivum and Nicotiana plumbaginifolia Viv. tissue cultures using the ethylene inhibitor AgNO3. Plant Cell Rep. 6:1–4

  21. Radke SE, Andrews BM, Moloney MM, Crouch ML, Kridl JC, Knauf VC (1988) Transformation of Brassica napus L. using Agrobacteriumtumefaciens: developmentally regulated expression of a reintroduced napin gene. Theor Appl Genet 75:685–694

  22. Sethi U, Basu A, Guha-Mukherjee S (1990) Control of cell proliferation and differentiation by modulators of ethylene biosynthesis and action in Brassica hypocotyl explants. Plant Sci 69:225–229

  23. Srivastava V, Reddy AS, Guha-Mukherjee (1988) Transformation and re-generation of Brassica oleracea mediated by an oncogenic Agrobacteriumtumefaciens. Plant Cell Rep 7:504–507

  24. Turner JC, Radke SE (1992) Inhibition of ethylene action increases transfor-mation rates of Brassica napus. In: S Knapp (ed) Proceedings of the Sev-enth Crucifer Genetics Workshop, Oregon State University, Corvallis, OR

Download references

Author information

Additional information

Communicated by C. F. Quiros

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Radke, S.E., Turner, J.C. & Facciotti, D. Transformation and regeneration of Brassica rapa using Agrobacterium tumefaciens . Plant Cell Reports 11, 499–505 (1992).

Download citation

Key words

  • Brassica
  • Oilseed rape
  • Gene transfer
  • Transgenic plants
  • Silver nitrate