Chromosome Transplantation and Applications of Flow Cytometry in Plants

  • A. M. M. de Laat
  • H. A. Verhoeven
  • K. Sree Ramulu
Part of the Biotechnology in Agriculture and Forestry book series (AGRICULTURE, volume 9)


Conventional plant breeding is gradually supplemented with novel techniques, e.g., in vitro propagation, selection at the cellular level and genetic manipulation. Genetic engineering has become realistic with the recent progress in molecular biology, cell biology, and tissue culture. By circumvention of sexual reproduction, the natural barriers of incompatibility can be bypassed. Whereas conventional breeding deals with combining whole genomes, differential transfer of genetic traits has become possible now. In the case where a monogenic trait has been characterized at the molecular level, the identified gene can be cloned and transferred into plant cells via DNA vectors (e.g., the Agrobacterium-derived Ti or Ri plasmids (Depicker et al. 1983) or viruses like CaMV (Brisson et al. 1984), or by direct DNA-uptake (Krens et al. 1982; Potrykus et al. 1985). The genetically modified cells are subsequently regenerated to transgenic plants.


Flow Cytometry Metaphase Chromosome Plant Chromosome Plant Protoplast Flow Sorting 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


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  1. Afonso CL, Harkins KR, Thomas-Compton MA, Krejci AE, Galbraith DW (1985) Selection of somatic hybrid plants in Nicotiana through fluorescence-activated sorting of protoplasts. Biotechnology 3: 811–816CrossRefGoogle Scholar
  2. Alexander RG, Cocking EC, Jackson PJ, Jett JH (1985) The characterization and isolation of plant heterokaryons by flow cytometry. Protoplasma 128: 52–58CrossRefGoogle Scholar
  3. Anderson LK, Stack SM, Fox MH, Chuanshan Z (1985) The relationship between genome size and synaptonemal complex length in higher plants. Exp Cell Res 156: 367–378PubMedCrossRefGoogle Scholar
  4. Bergounioux C, Perennes C, Miege C, Gadal P (1986) The effect of male sterility on protoplast division in Petunia hybrida. Cell cycle comparison by flow cytometry. Protoplasma 130: 138–144CrossRefGoogle Scholar
  5. Blumenthal AB, Dieden JD, Kapp LN, Sedat JW (1979) Rapid isolation of metaphase chromosomes containing high molecular weight DNA. J Cell Biol 81: 255–259PubMedCrossRefGoogle Scholar
  6. Brisson N, Paszkowski J, Penswick JR, Gronenborn B, Potrykus I, Mohn T (1984) Expression of a bacterial gene in plants by using a viral vector. Nature (Lond) 310: 511–514CrossRefGoogle Scholar
  7. Brown S, Renaudin JP, Prevot C, Guern J (1984) Flow cytometry and sorting of plant protoplasts: technical problems and physiological results from a study of pH and alkaloids in Catharanthus roseus. Physiol Veg 22: 541–554Google Scholar
  8. Brown S, Jullien M, Coutos-Thevenat P, Muller Ph, Renaudin JP (1986) Present developments of flow cytometry in plant biology. Biol Cell 58: 173–178Google Scholar
  9. Carlberg I, Glimelius K, Eriksson T (1984) Nuclear DNA content during the initiation of callus formation from isolated protoplasts of Solanum tuberosum L. Plant Sci Lett 35: 225–230CrossRefGoogle Scholar
  10. Conia J, Bergounioux C, Perennes C, Brown S, Muller Ph, Gadal P (1987) Chromosomes from protoplasts of Petunia hybrids, flow analysis and sorting. Biol Cell 58 Suppl: 5aGoogle Scholar
  11. Coyle MB, Strauss B (1970) Cell killing and the accumulation of breaks in the DNA of HEp-2 cells incubated in the presence of hydroxyurea. Cancer Res 30: 2314–2319PubMedGoogle Scholar
  12. Davies KE, Young BD, Elles RG, Hill ME, Williamson R (1981) Cloning of a representative genomic library of the human X-chromosome after sorting by flow cytometry. Nature 293: 374–378PubMedCrossRefGoogle Scholar
  13. de Laat AMM, Blaas J (1983) Isolation, purification and sorting by flow cytometry of metaphase chromosomes of Haplopappus gracilis. In: Potrykus I, Harms CT, Hinnen A, Hütter R, King PJ, Shillito A (eds) 6th Int Protoplast Symp, Poster Proc. Birkhäuser, Basel, pp 120–121Google Scholar
  14. de Laat AMM, Blaas J (1984) Flow cytometric characterization and sorting of plant chromosomes. Theor Appl Genet 67: 463–467CrossRefGoogle Scholar
  15. de Laat AMM, Blaas J (1987) An improved method for protoplast micro-injection suitable for transfer of entire plant chromosomes. Plant Sci 50: 161–169CrossRefGoogle Scholar
  16. de Laat AMM, Shell JHN (1986) Integrity of metaphase chromosomes of Haplopappus gracilis ( Nutt.) Gray isolated by flow cytometry. Plant Sci 47: 145–151CrossRefGoogle Scholar
  17. de Laat AMM, Verhoeven HA, Sree Ramulu K, Dijkhuis P (1987) Efficient induction by amiprophosmethyl and flow cytometric sorting of micronuclei in Nicotiana plumbaginifolia. Planta 172: 473–478CrossRefGoogle Scholar
  18. Depicker A, Montagu van M, Shell J (1983) Plant cell transformation by Agrobacterium plasmids. In: Kosuge T, Meredith CP, Hollaender A (eds) Genetic engineering of plants: an agricultural perspective. Plenum, New York, pp 143–176Google Scholar
  19. Dudits D, Praznovzky T (1985) Intergeneric gene transfer by protoplast fusion and uptake of isolated chromosomes. In: Zaitlin M, Day P, Hollaender A (eds) Biotechnology in plant science: relevance to agriculture in the eighties. Academic Press, New York London Orlando, pp 115–128Google Scholar
  20. Eriksson T (1966) Partial synchronization of cell division in suspension cells of Haplopappus gracilis. Physiol Plant 19: 900–910CrossRefGoogle Scholar
  21. Famelaer Y, Cammaerts D, Karp A, Sidorov V, De Brouwer D, Negrutiu I, Jacobs M (1986) Cellular engineering by ‘gamma fusion’ and ‘egg transformation’: Recent experimental data and applications in plant breeding. In: Proc Symp Nuclear techniques and in vitro culture for plant improvement, IAEA, Vienna, pp 453–460Google Scholar
  22. Fournier REK (1982) Microcell-mediated chromosome transfer. In: Shay J (ed) Techniques in somatic cell genetics. Plenum, New York, pp 309–327Google Scholar
  23. Galbraith DW (1984) Flow cytometric analysis of the cell cycle. In: Vasil IK (ed) Cell culture and somatic cell genetics of plants, vol 1. Academic Press, New York London, pp 765–777Google Scholar
  24. Galbraith DW, Shields BA (1982) The effect of inhibitors of cell wall synthesis on tobacco protoplast development. Physiol Plant 55: 25–30CrossRefGoogle Scholar
  25. Galbraith DW, Harkins KR, Maddox JM, Ayres NM, Sharma DP, Firoozabady E (1983) Rapid flow cytometric analysis of the cell cycle in intact plant tissues. Science 220: 1049–1051PubMedCrossRefGoogle Scholar
  26. Glimelius K (1987) Enrichment of somatic hybrids with flow cytometry. IAPTC Newslett 52: 2–10Google Scholar
  27. Glimelius K, Djupsjöbacka M, Fellner-Feldegg H (1986) Selection and enrichment of plant protoplasts heterokaryons of Brassicaceae by flow sorting. Plant Sci 45: 133–141CrossRefGoogle Scholar
  28. Griesbach RJ (1987) Chromosome-mediated transformation via microinjection. Plant Sci 50: 69–77CrossRefGoogle Scholar
  29. Griesbach RJ, Malmberg RL, Carlson PS (1982) An improved technique for the isolation of higher plant chromosomes. Plant Sci Lett 24: 55–60CrossRefGoogle Scholar
  30. Hadlaczky G, Sumner AT, Ross A (1981) Protein-depleted chromosomes. I. Structure of isolated protein-depleted chromosomes. Chromosoma 81: 537–555PubMedCrossRefGoogle Scholar
  31. Hadlaczky G, Bisztray G, Praznovszky T, Dudits D (1983) Mass isolation for plant chromosomes and nuclei. Planta 157: 278–285CrossRefGoogle Scholar
  32. Hammer RE, Pursel VG, Rexroad CE, Wall RJ, Bolt DJ, Ebert KM, Palmiter RD, Brinster RL (1985) Production of transgenic rabbits, sheeps and pigs by microinjection. Nature (Lond) 315: 680–683CrossRefGoogle Scholar
  33. Hanisch ten Cate CH, Sree Ramulu K (1987) Callus growth, tumour development and polyploidization in the tetraploid potato cultivar Bintje. Plant Sci 49: 209–216CrossRefGoogle Scholar
  34. Harkins KR, Galbraith DW (1984) Flow sorting and culture of plant protoplasts. Physiol Plant 60: 43–52CrossRefGoogle Scholar
  35. Harkins KR, Galbraith DW (1987) Factors governing the flow cytometric analysis and sorting of large biological particles. Cytometry 8: 60–70PubMedCrossRefGoogle Scholar
  36. Hepher A, Gates P, Boulter D (1985) Microinjection of DNA into pollen, ovaries, and somatic cells. In: EC contractants Meet Biomolecular engineering programme, Copenhagen, Oct 9–10. Abstr, pp 30–31Google Scholar
  37. Jackson RC (1957) New low chromosome number for plants. Sci 126: 1115–1116CrossRefGoogle Scholar
  38. Jett JH, Alexander RG (1985) Droplet sorting of large particles. Cytometry 6: 484–486PubMedCrossRefGoogle Scholar
  39. Karlsson SB, Vasil IK (1986) Growth, cytology and flow cytometry of embryogenic cell suspension cultures of Panicum maximum Jacq. and Pennisetum purpureum Schum. J Plant Physiol 123: 211–227Google Scholar
  40. Krens FA, Molendijk L, Wullems GJ, Schilperoort RA (1982) In vitro transformation of plant protoplasts with Ti-plasmid DNA. Nature (Lund) 296: 72–74CrossRefGoogle Scholar
  41. Larkin PJ, Scowcroft WR (1981) Somaclonal variation — a novel source of variability from cell cultures for plant improvement. Theor Appl Genet 60: 197–214CrossRefGoogle Scholar
  42. Lörz H, Potrykus I (1978) Investigations on the transfer of isolated nuclei into plant protoplasts. Theor Appl Genet 53: 251–256Google Scholar
  43. Lugo TG, Fournier REK (1986) Microcell fusion and mammalian gene transfer. In: Kucherlapati R (ed) Gene transfer. Plenum, New York, pp 79–93Google Scholar
  44. Mathias RJ, Boyd LA, Gleddie SA, Flavell RB (1986) Transformation of wheat by microinjection. In: EC contractants Meet Biomolecular engineering programme, Wageningen, Jan 27–30. Abstr, pp 99–100Google Scholar
  45. Matthews BF (1983) Isolation of mitotic chromosomes from partially synchronized carrot (Daucus carota) cell suspension cultures. Plant Sci Lett 31: 165–172CrossRefGoogle Scholar
  46. Mce>Sree Ramulu OW, Peterson JL (1980) Chromosome-mediated gene transfer in mammalian cells. Annu Rev Genet 14: 321–345PubMedCrossRefGoogle Scholar
  47. Meadows MG (1982) Characterization of cells and protoplasts of the B73 maize cell line. Plant Sci Lett 28: 337–348Google Scholar
  48. Melamed MR, Mullaney PF, Mendelsohn ML (1979) Flow cytometry and sorting. John Wiley & Sons, New YorkGoogle Scholar
  49. Muirhead KA, Hovan PK (1984) Applications of flow cytometry to tissue culture systems. In: Maramorosch K (ed) Advances in cell culture, vol 3. Academic Press, New York London, pp 57–91Google Scholar
  50. Petit P, Conia J, Brown SC, Bergounioux C (1986) Cytométrie en flux et biolotechnologies végétales. Un avenir prometteur. Biofutur 51: 128–139Google Scholar
  51. Pinkel D (1981) On the possibility of automated scoring of pollen mutants. Environ Health Perspectives 37: 133–136CrossRefGoogle Scholar
  52. Potrykus I, Shillito RD, Saul MW, Paszkowski J (1985) Direct gene transfer. State of art and future potential. Plant Mol Biol Rep 3: 117–128CrossRefGoogle Scholar
  53. Puite KJ, Ten Broeke WRR (1983) DNA staining of fixed and non-fixed plant protoplasts for flow cytometry with Hoechst 33342. Plant Sci Lett 32: 79–88CrossRefGoogle Scholar
  54. Puite KJ, Gilissen LJW, de Laat AMM (1983) First steps to chromosome transplantation with Hapopappus gracilis using flow cytometry. In: Potrykus I, Harms CT, Hinnen A, Hütter R, King PJ, Shillito A (eds) 6th Int Protoplast Symp Poster Proc. Birkhäuser, Basel, pp 122–123Google Scholar
  55. Saxena PK, Mii M, Crosby WL, Fowke LC, King J (1986) Transplantation of isolated nuclei into plant protoplasts. A novel technique for introducing foreign DNA into plant cells. Planta 168: 29–35CrossRefGoogle Scholar
  56. Sharma DP, Firoozabady E, Ayres NM, Galbraith DW (1983) Improvement of anther culture in Nicotiana: Media, cultural conditions and flow cytometric determination of ploidy levels. Z Planzenphysiol 111: 441–451Google Scholar
  57. Sree Ramulu K (1986) Case histories of genetic variability in vitro: Potato. In: Vasil IK (ed) Cell culture and somatic cell genetics of plants, vol 3. Academic Press, New York London, pp 449–473Google Scholar
  58. Sree Ramulu K, Dijkhuis P (1986) Flow cytometric analysis of polysomaty and in vitro genetic instability in potato. Plant Cell Rep 3: 234–237Google Scholar
  59. Sree Ramulu K, Verhoeven HA, Dijkhuis P (1988) Mitotic dynamics of micronuclei induced by amiprophos-methyl and prospects for chromosome-mediated gene transfer in plants. Theor Appl Genet 75: 575–584CrossRefGoogle Scholar
  60. Sundberg E, Glimelius K (1986) A method for production of interspecific hybrids within Brassicaceae via somatic hybridization, using resynthesis of Brassica napus as a model. Plant Sci 43: 155–162CrossRefGoogle Scholar
  61. Szabados L, Hadlaczky G, Dudits D (1981) Uptake of isolated plant chromosomes by plant protoplasts. Planta 151: 141–145CrossRefGoogle Scholar
  62. Valk HCPM van der, Verhoeven HA (1988) Application of flow cytometry for the isolation of metaphase chromosomes from protoplasts of Nicotiana plumbaginifolia. In: 7th Int Protoplast Symp, Wageningen. Nijhoff, Dordrecht (in press)Google Scholar
  63. Verhoeven HA, de Laat AMM, Blaas J (1986) Methodological developments for chromosome-mediated gene transfer in higher plants. In: Magnien E (ed) Biomolecular engineering in the European Community, achievements of the research programme (1982–1986). Final Rep, Nijhoff, Dordrecht, pp 977–981Google Scholar
  64. Verhoeven HA, de Laat AMM, Sree Ramulu K, Dijkhuis P, Blaas J, Vloten-Doting L van (1987) Induction, isolation and flow cytometric sorting of metaphase chromosomes and micronuclei from cell suspension cultures of Nicotiana plumbaginifolia. EC contractant Meeting Biotechnology action programme, Louvain La Neuve, March. Abstr, pp 119–120Google Scholar
  65. Willadsen SM (1986) Nuclear transplantation in sheep embryos. Nature (Lond) 320: 63–65CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1989

Authors and Affiliations

  • A. M. M. de Laat
    • 1
  • H. A. Verhoeven
    • 1
  • K. Sree Ramulu
    • 1
  1. 1.Research Institute ItalWageningenThe Netherlands

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