Summary
32P-labelled probes prepared by nick-translation, two methods (A and B) of labelling PSTV cDNA in M13 phage DNA, and RNA transcription were compared for their sensitivity in detecting PSTV. Nick-translated and method B probes were the least sensitive. The minimum concentration of PSTV detected by these probes was 100 pg. Method A probe detected 10–20 pg of PSTV whereas RNA transcripts were capable of detecting as little as 0.33 pg of PSTV. RNA transcripts are easy to prepare, and their use in hybridization analysis do not cause background reactions. Sometimes low non-specific signals are produced but they can be eliminated by washing membranes with RNase A. RNA probes are now used for routine detection of PSTV in tuber flesh and sprouts, leaves and true seed of potato at the International Potato Center (CIP).
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References
Barker, J. M., J. L. McInnes, P. J. Murphy & R. H. Symons, 1985. Dot-blot procedure with (32P) DNA probes for the sensitive detection of Avocado sunblotch and other viroids in plants.Journal of Virological Methods 10: 87–98.
Cress, D. E., M. C. Kiefer & R. A. Owens, 1983. Construction of infectious potato spindle tuber viroid cDNA.Nucleic Acids Research 11: 6821–6835.
Diener, T. O., 1979. Viroids and viroid diseases. John Wiley & Sons, New York. 252 pp.
Harris, P. S. & I. A. Browning, 1980. The effects of temperature and light on the symptom expression and viroid concentration in tomato of a severe strain of potato spindle tuber viroid.Potato Research 23: 85–93.
Maniatis, T., E. F. Fritsch & J. Sambrook, 1982. Molecular Cloning, a laboratory manual. Cold Spring Harbor Laboratory, New York, 545 pp.
Melton, D. A., P. A. Krieg, M. R. Rebagliati, T. Maniatis, K. Zinn & M. R. Green, 1984. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter.Nucleic Acids Research 12: 7035–7056.
Meinkoth, J. & G. Wahl, 1984. Hybridization of nucleic acids immobilized on solid support.Analytical Biochemistry 138: 267–284.
Morris, T. J. & N. S. Wright, 1975. Detection on polyacrylamide gel of a diagnostic nucleic acid from tissue infected with potato spindle tuber viroid.American Potato Journal 52: 57–63.
Owens, R. A. & T. O. Diener, 1981. Sensitive and rapid diagnosis of potato spindle tuber viroid disease by nucleic acid hybridization.Science 213: 670–672.
Palukaitis, P., S. Cotts & M. Zaitlin, 1985. Detection and identification of viroids and viral nucleic acids by ‘dot blot’ hybridization.Acta Horticulturae 164: 109–118.
Raymer, W. B. & M. J. O'Brien 1962. Transmission of potato spindle tuber virus to tomato.American Potato Journal 39: 401–408.
Salazar, L. F., R. A. Owens, D. R. Smith & T. O. Diener, 1983. Detection of potato spindle tuber viroid by nucleic acid spot hybridization: Evaluation with tuber sprouts and true potato seed.American Potato Journal 60: 587–597.
Salazar, L. F., L. Schilde-Rentschler & R. Lizárraga, 1985. Elimination of potato spindle tuber viroid from potato by cold treatment and meristem culture. In: Maramorosch, K. & J. J. McKelvey Jr. (Eds.), Subviral pathogens of plants and animals: viroids and prions. Academic Press. p. 137–150.
Van Gelder, W. M. J. & A. Treur, 1982. Testing of imported potato genotypes for potato spindle tuber viroid with a tomato-intermediate/electrophoresis combined method.EPPO Bulletin 12: 297–305.
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Salazar, L.F., Balbo, I. & Owens, R.A. Comparison of four radioactive probes for the diagnosis of potato spindle tuber viroid by nucleic acid spot hybridization. Potato Res 31, 431–442 (1988). https://doi.org/10.1007/BF02357880
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DOI: https://doi.org/10.1007/BF02357880