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RNA interference-based resistance in transgenic tomato plants against Tomato yellow leaf curl virus-Oman (TYLCV-OM) and its associated betasatellite.

Ammara Ue, Mansoor S, Saeed M, Amin I, Briddon RW, Al-Sadi AM - Virol. J. (2015)

Bottom Line: Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite).These results show that RNAi can be used to develop resistance against geminiviruses in tomato.Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.

View Article: PubMed Central - PubMed

Affiliation: Department of Crop Sciences, College of Agriculture and Marine Sciences, Sultan Qaboos University, P.O. Box-34, 123, Al-Khod, Oman. ammarajawad@gmail.com.

ABSTRACT

Background: Tomato yellow leaf curl virus (TYLCV), a monopartite begomovirus (family Geminiviridae) is responsible for heavy yield losses for tomato production around the globe. In Oman at least five distinct begomoviruses cause disease in tomato, including TYLCV. Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite). RNA interference (RNAi) can be used to develop resistance against begomoviruses at either the transcriptional or post-transcriptional levels.

Results: A hairpin RNAi (hpRNAi) construct to express double-stranded RNA homologous to sequences of the intergenic region, coat protein gene, V2 gene and replication-associated gene of Tomato yellow leaf curl virus-Oman (TYLCV-OM) was produced. Initially, transient expression of the hpRNAi construct at the site of virus inoculation was shown to reduce the number of plants developing symptoms when inoculated with either TYLCV-OM or TYLCV-OM with ToLCB-OM to Nicotiana benthamiana or tomato. Solanum lycopersicum L. cv. Pusa Ruby was transformed with the hpRNAi construct and nine confirmed transgenic lines were obtained and challenged with TYLCV-OM and ToLCB-OM by Agrobacterium-mediated inoculation. For all but one line, for which all plants remained symptomless, inoculation with TYLCV-OM led to a proportion (≤25%) of tomato plants developing symptoms of infection. For inoculation with TYLCV-OM and ToLCB-OM all lines showed a proportion of plants (≤45%) symptomatic. However, for all infected transgenic plants the symptoms were milder and virus titre in plants was lower than in infected non-transgenic tomato plants.

Conclusions: These results show that RNAi can be used to develop resistance against geminiviruses in tomato. The resistance in this case is not immunity but does reduce the severity of infections and virus titer. Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.

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Related in: MedlinePlus

Southern blot analysis of transgenic tomato plants harboring the hpRNAi construct inoculated with TYLCV-OM probed for the presence of TYLCV-OM (A) and inoculated with TYLCV-OM/ToLCB-OM probed for the presence of ToLCB-OM (B). The DNA samples run on the gel were extracted from a non-transgenic non-inoculated tomato cv. Pusa Ruby plant (N), symptomatic TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B) inoculated non-transgenic Pusa Ruby plants (C) and two plants each of 9 transgenic Pusa Ruby lines (as indicated on the figure) inoculated with either TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B). Approx. 10 μg of total DNA was loaded on each case and the samples were extracted at 60 dpi. The positions of the viral single-stranded (ss) and supercoiled (sc) replicative DNA forms are indicated.
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Fig4: Southern blot analysis of transgenic tomato plants harboring the hpRNAi construct inoculated with TYLCV-OM probed for the presence of TYLCV-OM (A) and inoculated with TYLCV-OM/ToLCB-OM probed for the presence of ToLCB-OM (B). The DNA samples run on the gel were extracted from a non-transgenic non-inoculated tomato cv. Pusa Ruby plant (N), symptomatic TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B) inoculated non-transgenic Pusa Ruby plants (C) and two plants each of 9 transgenic Pusa Ruby lines (as indicated on the figure) inoculated with either TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B). Approx. 10 μg of total DNA was loaded on each case and the samples were extracted at 60 dpi. The positions of the viral single-stranded (ss) and supercoiled (sc) replicative DNA forms are indicated.

Mentions: A Southern blot hybridization analysis of inoculated tomato plants detected high levels of both virus and betasatellite in symptomatic non-transgenic plants. However, with the exception of one plant, in which low levels of the betasatellite were detected, no virus or betasatellite DNA could be detected in inoculated transgenic plants (Figure 4).Figure 4


RNA interference-based resistance in transgenic tomato plants against Tomato yellow leaf curl virus-Oman (TYLCV-OM) and its associated betasatellite.

Ammara Ue, Mansoor S, Saeed M, Amin I, Briddon RW, Al-Sadi AM - Virol. J. (2015)

Southern blot analysis of transgenic tomato plants harboring the hpRNAi construct inoculated with TYLCV-OM probed for the presence of TYLCV-OM (A) and inoculated with TYLCV-OM/ToLCB-OM probed for the presence of ToLCB-OM (B). The DNA samples run on the gel were extracted from a non-transgenic non-inoculated tomato cv. Pusa Ruby plant (N), symptomatic TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B) inoculated non-transgenic Pusa Ruby plants (C) and two plants each of 9 transgenic Pusa Ruby lines (as indicated on the figure) inoculated with either TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B). Approx. 10 μg of total DNA was loaded on each case and the samples were extracted at 60 dpi. The positions of the viral single-stranded (ss) and supercoiled (sc) replicative DNA forms are indicated.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4359554&req=5

Fig4: Southern blot analysis of transgenic tomato plants harboring the hpRNAi construct inoculated with TYLCV-OM probed for the presence of TYLCV-OM (A) and inoculated with TYLCV-OM/ToLCB-OM probed for the presence of ToLCB-OM (B). The DNA samples run on the gel were extracted from a non-transgenic non-inoculated tomato cv. Pusa Ruby plant (N), symptomatic TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B) inoculated non-transgenic Pusa Ruby plants (C) and two plants each of 9 transgenic Pusa Ruby lines (as indicated on the figure) inoculated with either TYLCV-OM (panel A) or TYLCV-OM/ToLCB-OM (panel B). Approx. 10 μg of total DNA was loaded on each case and the samples were extracted at 60 dpi. The positions of the viral single-stranded (ss) and supercoiled (sc) replicative DNA forms are indicated.
Mentions: A Southern blot hybridization analysis of inoculated tomato plants detected high levels of both virus and betasatellite in symptomatic non-transgenic plants. However, with the exception of one plant, in which low levels of the betasatellite were detected, no virus or betasatellite DNA could be detected in inoculated transgenic plants (Figure 4).Figure 4

Bottom Line: Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite).These results show that RNAi can be used to develop resistance against geminiviruses in tomato.Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.

View Article: PubMed Central - PubMed

Affiliation: Department of Crop Sciences, College of Agriculture and Marine Sciences, Sultan Qaboos University, P.O. Box-34, 123, Al-Khod, Oman. ammarajawad@gmail.com.

ABSTRACT

Background: Tomato yellow leaf curl virus (TYLCV), a monopartite begomovirus (family Geminiviridae) is responsible for heavy yield losses for tomato production around the globe. In Oman at least five distinct begomoviruses cause disease in tomato, including TYLCV. Unusually, TYLCV infections in Oman are sometimes associated with a betasatellite (Tomato leaf curl betasatellite [ToLCB]; a symptom modulating satellite). RNA interference (RNAi) can be used to develop resistance against begomoviruses at either the transcriptional or post-transcriptional levels.

Results: A hairpin RNAi (hpRNAi) construct to express double-stranded RNA homologous to sequences of the intergenic region, coat protein gene, V2 gene and replication-associated gene of Tomato yellow leaf curl virus-Oman (TYLCV-OM) was produced. Initially, transient expression of the hpRNAi construct at the site of virus inoculation was shown to reduce the number of plants developing symptoms when inoculated with either TYLCV-OM or TYLCV-OM with ToLCB-OM to Nicotiana benthamiana or tomato. Solanum lycopersicum L. cv. Pusa Ruby was transformed with the hpRNAi construct and nine confirmed transgenic lines were obtained and challenged with TYLCV-OM and ToLCB-OM by Agrobacterium-mediated inoculation. For all but one line, for which all plants remained symptomless, inoculation with TYLCV-OM led to a proportion (≤25%) of tomato plants developing symptoms of infection. For inoculation with TYLCV-OM and ToLCB-OM all lines showed a proportion of plants (≤45%) symptomatic. However, for all infected transgenic plants the symptoms were milder and virus titre in plants was lower than in infected non-transgenic tomato plants.

Conclusions: These results show that RNAi can be used to develop resistance against geminiviruses in tomato. The resistance in this case is not immunity but does reduce the severity of infections and virus titer. Also, the betasatellite may compromise resistance, increasing the proportion of plants which ultimately show symptoms.

Show MeSH
Related in: MedlinePlus