1887

Abstract

We describe here a convenient and efficient system for studying turnip yellow mosaic virus (TYMV) replication in leaf protoplasts. Inoculation of rapeseed ( ) or Chinese cabbage ( ) protoplasts was achieved via electroporation, and sensitive detection of viral RNA products was performed by Northern blot analyses using a non-radioactive digoxigenin-labelled cDNA probe. Virus replication was detected when 1.5×10 rapeseed protoplasts were inoculated with 20 ng of TYMV RNA. Electrotransfection of TYMV RNA was more efficient in rapeseed than in Chinese cabbage protoplasts, and gave somewhat higher signals than those of TYMV virions. TYMV RNA appeared to replicate equally well whether the protoplasts were incubated in the dark or under constant light.

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1993-09-01
2024-04-19
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References

  1. Balint R, Cohen SS. 1985; The effects of dicyclohexylamine on polyamine biosynthesis and incorporation into turnip yellow mosaicvirus in Chinese cabbage protoplasts infected in vitro.. Virology 144:194–203
    [Google Scholar]
  2. Benicourt C, Pere JP, Haenni AL. 1978; Translation of TYMV RNA into high molecular weight proteins.. FEBS Letters 86:268–272
    [Google Scholar]
  3. Bronstein I, Voyta JC, Lazzari KG, Murphy O, Edwards B, Kricka LJ. 1990; Rapid and sensitive detection of DNA insouthern blots with chemiluminescence.. Bio/Techniques 8:310–314
    [Google Scholar]
  4. Candresse T, Macquaire G, Brault V, Monsion M, Dunez J. 1990; 32P- and biotin-labelled in vitro transcribed cRNA probesfor the detection of potato spindle tuber viroid and chrysanthemumstunt viroid.. Research in Virology 141:97–107
    [Google Scholar]
  5. Dean C, van den Elzen P, Tamaki S, Dunsmuir P, Bedbrook J. 1985; Differential expression of the eight genes of the petuniaribulose bisphosphate carboxylase small subunit multi-gene family.. EMBO Journal 4:3055–3061
    [Google Scholar]
  6. Domier LL, Franklin KM, Hunt AG, Rhoads RE, Shaw JG. 1989; Infectious in vitro transcripts from cloned cDNAof a potyvirus, tobacco vein mottling virus.. Proceedings of theNational Academy of Sciences, U.S.A. 86:3509–3513
    [Google Scholar]
  7. Fernandez-Gonzalez O, Renaudin J, Bove JM. 1980; Infection of chlorophyll-less protoplasts from etiolated Chinesecabbage hypocotyls by turnip yellow mosaic virus.. Virology 104:262–265
    [Google Scholar]
  8. Fraser L, Matthews REF. 1979; Strain-specific pathways of cytological change in individual Chinese cabbage protoplasts infectedwith turnip yellow mosaic virus.. Journal of General Virology 45:623–630
    [Google Scholar]
  9. Gamborg OL, Miller RA, Ojima K. 1968; Nutrient requirements of suspension cultures of soybean root cells.. Experimental Cell Research 50:151–158
    [Google Scholar]
  10. Guerche P, Charbonnier L, Jouanin L, Tourneur C, Paszkowski J, Pelletier G. 1987a; Direct gene transfer byelectroporation in Brassica napus.. Plant Science 52:111–116
    [Google Scholar]
  11. Guerche P, Bellini C, Le Moullec JM, Caboche M. 1987b; Use of a transient expression assay for the optimization of directgene transfer into tobacco mesophyll protoplasts by electroporation.. Biochimie 69:621–628
    [Google Scholar]
  12. Hibi T. 1989; Electrotransfection of plant protoplasts with viral nucleic acids.. Advances in Virus Research 37:329–342
    [Google Scholar]
  13. Holness CL, Lomonossoff GP, Evans D, Maule AJ. 1989; Identification of the initiation codons for translation ofcowpea mosaic virus middle component RNA using site-directedmutagenesis of an infectious clone.. Virology 172:311–320
    [Google Scholar]
  14. Leberman R. 1966; The isolation of plant viruses by means of ‘simple’ coacervates.. Virology 30:341–347
    [Google Scholar]
  15. McMaster GK, Carmichael GG. 1977; Analysis of single-and double-stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange.. Proceedings of theNational Academy of Sciences, U.S.A. 74:4835–4838
    [Google Scholar]
  16. Matsunaga R, Sawamura K, De Kok M, Makino T, Miki K, Kojima M, Tsuchizaki T, Hibi T. 1992; Electrotransfection ofprotoplasts from tomato, wild tomato, barley and chrysanthemumwith tobacco mosaic virus RNA.. Journal of General Virology 73:763–766
    [Google Scholar]
  17. Matthews REF. 1991 Plant Virology, 3rd edn.. New York: Academic Press.;
    [Google Scholar]
  18. Melton DA, Krieg PA, Rebagliati MR, Maniatis T, Zinn K, Green MR. 1984; Efficient in vitro synthesis of biologicallyactive RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter.. Nucleic Acids Research 12:7035–7056
    [Google Scholar]
  19. Morch MD, Boyer JC, Haenni AL. 1988; Overlapping open reading frames revealed by complete nucleotide sequencing ofturnip yellow mosaic virus genomic RNA.. Nucleic Acids Research 16:6157–6173
    [Google Scholar]
  20. Nishiguchi M, Sato T, Motoyoshi F. 1987; An improved method for electroporation in plant protoplasts: infection of tobaccoprotoplasts by tobacco mosaic virus particles.. Plant Cell Reports 6:90–93
    [Google Scholar]
  21. Rao ALN, Huntley CC, Marsh LE, Hall TC. 1990; Analysis of RNA stability and (—) strand content in viral infectionsusing biotinylated RNA probes.. Journal of Virological Methods 30:239–250
    [Google Scholar]
  22. Renaudin J, Bove JM, Otsuki Y, Takebe I. 1975; Infection of Brassica leaf protoplasts by turnip yellow mosaic virus.. Molecularand General Genetics 141:59–68
    [Google Scholar]
  23. Rouan D. 1991 Transf ert direct de gbnes chez les Brassica: application 7l la rdsistance au virus de la mosa?que du chou-fleur. Ph.D. thesis, Universit6 Paul Sabatier: Toulouse.;
    [Google Scholar]
  24. Rouan D, Guerche P. 1991; Transformation and regeneration of oilseed rape protoplasts.. Plant Tissue Culture ManualEdited by Lindsey K. The Netherlands: Kluwer Academic Publishers.;
    [Google Scholar]
  25. Samac DA, Nelson SE, LoeschFries LS. 1983; Virus protein synthesis in alfalfa mosaic virus infected alfalfa protoplasts.. Virology 131:455–462
    [Google Scholar]
  26. Sugimura Y, Matthews REF. 1981; Timing of the synthesis of empty shells and minor nucleoproteins in relation to turnip yellowmosaic virus synthesis in Brassica protoplasts.. Virology 112:70–80
    [Google Scholar]
  27. Veidt I, Bouzoubaa SE, Leiser RM, ZieglerGraff V, Guilley H, Richards K, Jonard G. 1992; Synthesis of full-length transcripts of beet western yellows virus RNA: messengerproperties and biological activity in protoplasts.. Virology 186:192–200
    [Google Scholar]
  28. Viry M, Serghini MA, Hans F, Ritzenthaler C, Pinck M, Pinck L. 1993; Biologically active transcripts from cloned cDNAof genomic grapevine fanleaf nepovirus RNAs.. Journal of GeneralVirology 74:169–174
    [Google Scholar]
  29. Watanabe Y, Meshi T, Okada Y. 1987; Infection of tobacco protoplasts with in vitro transcribed tobacco mosaic virus RNAusing an improved electroporation method.. FEBS Letters 219:65–69
    [Google Scholar]
  30. Watts JW, King JM, Stacey NJ. 1987; Inoculation of protoplasts with viruses by electroporation.. Virology 157:40–46
    [Google Scholar]
  31. Weiland JJ, Dreher TW. 1989; Infectious TYMV RNA from cloned cDNA: effects in vitro and in vivo of point substitutions in theinitiation codons of two extensively overlapping ORFs.. NucleicAcids Research 17:4675–4687
    [Google Scholar]
  32. Wilkinson M, Doskow J, Lindsey S. 1991; RNA blots: staining procedures and optimization of conditions.. Nucleic Acids Research 19:679
    [Google Scholar]
  33. Young MJ, Kelly L, Larkin PJ, Waterhouse PM, GerlaCH WL. 1991; Infectious in vitro transcripts from a cloned cDNA of barley yellow dwarf virus.. Virology 180:372–379
    [Google Scholar]
  34. Zaccomer B, Cellier F, Boyer JC, Haenni AL, Tepfer M. 1993; Transgenic plants that express genes including the 3′ untranslated region of the turnip yellow mosaic virus (TYMV)genome are partially protected against TYMV infection.. Gene (inpress)
    [Google Scholar]
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