Induced Systemic Resistance in Potato (Solanum tuberosum L.) Following Localised Potato Virus X Infection

Barrett, Stuart James (1991) Induced Systemic Resistance in Potato (Solanum tuberosum L.) Following Localised Potato Virus X Infection. PhD thesis, University of Glasgow.

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Abstract

The aim of this project was to investigate the resistance response of potato to potato virus X (PVX) and the possible correlation between resistance and the occurrence of induced systemic resistance (inferring greater resistance to a second challenge from PVX) at a molecular level. The investigation used a number of methods to look at the changes in gene expression which accompanied the invasion of PVX in a restricting potato cultivar. The objective was to identify genes involved in PVX resistance and to characterise them; looking at their expression in comparison to healthy and mock-inoculated controls. Group 2 strains of PVX, when inoculated onto potato cultivars carrying the Nb gene, form local lesions but do not spread systemically. In comparison. Group 3 strains of PVX are able to infect such plants systemically (Cockerham, 1970). Cultivar Pentland Ivory, which carries the Nb gene, was inoculated with the Group 2 PVX strain KEBRS and developed local lesions after six days. When non-inoculated upper leaves of these plants were inoculated with PVX-KEBRS, or a Group 3 PVX strain (Gp3RS), 20 days after the first inoculation, the plants expressed a greater level of resistance than control plants which had not been earlier inoculated. Reduced lesion number on plants challenged with a Group 2 virus strain, or a reduced systemic infection when challenged by a Group 3 virus strain, indicated increased resistance. However, the level of resistance induced by pre-inoculation with PVX-KEBRS in comparison to mock-inoculation with water did not show a statistically significant difference. RNA was extracted from PVX-KEBRS-inoculated, mock-inoculated (water) and healthy uninoculated (control) plants, from both the inoculated and upper leaves of Pentland Ivory plants, at various times after inoculation. The RNA was translated in vitro. The polypeptides were separated using one and two-dimensional PAGE systems. The results obtained from one-dimensional gels showed that significant changes were first observed 7 days after inoculation, both locally and systemically to the point of inoculation with PVX-KEBRS, compared to both the control treatments. Using two-dimensional gels the in vitro RNA translation polypeptides (from RNA samples taken 7, 8, 9 and 12 days after inoculation) were further resolved. This identified that most mRNA induced changes in PVX-KEBRS-inoculated leaves were seen 7-8 days after inoculation while the greatest number of systemic changes were seen 9-12 days after inoculation. There were also several mRNAs which were induced both locally and systemically by PVX-KEBRS-inoculation and mock- inoculation. Thus inoculation of Pentland Ivory plants with PVX-KEBRS, or merely mock inoculating them with water, induced the synthesis of several mRNAs both locally and systemically to the inoculated leaf. Poly A+ RNA was extracted from non-inoculated leaves of Pentland Ivory plants, 12 days after inoculation with PVX-KEBRS, and used to construct a cDNA library in the lambda vector NM1149, The library was screened with a PR-1a and a thaumatin-like cDNA from tobacco and this identified mRNA showing homology to the two clones at a frequency of 0.1% and 0.05% respectively. Twenty-eight thousand plaques from the library were screened with a combination of subtractive and differential ss cDNA probes made from PVX-KEBRS-inoculated, mock- inoculated and healthy plants from the non-inoculated leaves 12 days after inoculation. This screening resulted in the identification of six clones which were induced systemically by PVX-KEBRS-inoculation. Three of these six clones were further investigated using Northern blot analysis. This identified a clone which was strongly induced by either PVX-KEBRS or mock-inoculation in the inoculated leaf. A second clone was systemically induced by PVX-KEBRS inoculation, while a third clone was found to be poorly differentially expressed between treatments. Therefore, the clones investigated so far have identified a range of mRNAs which are induced locally or systemically by PVX-KEBRS or mock-inoculation. These may be involved in the phenomena of induced systemic resistance.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Additional Information: Adviser: D D Clarke
Keywords: Plant sciences, Plant pathology
Date of Award: 1991
Depositing User: Enlighten Team
Unique ID: glathesis:1991-75353
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 19 Nov 2019 20:26
Last Modified: 19 Nov 2019 20:26
URI: http://theses.gla.ac.uk/id/eprint/75353

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