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Plant Cell Reports

, Volume 28, Issue 4, pp 639–647 | Cite as

Quantitative evaluation of six different viral suppressors of silencing using image analysis of transient GFP expression

  • Taniya Dhillon
  • Joseph M. Chiera
  • John A. Lindbo
  • John J. FinerEmail author
Genetic Transformation and Hybridization

Abstract

The effects of six different plant viral suppressors of gene silencing were compared using an automated image collection and analysis system developed for continual monitoring of GFP expression. Suppressors were introduced into lima bean cotyledonary tissues either as 3′-GFP translational fusions or as co-introductions with the GFP gene on a separate plasmid. The resultant transient expression profiles for each suppressor depended on whether the suppressor was introduced as a fusion or co-introduced on separate plasmids. As co-introductions, the silencing suppressors HCPro (from Tobacco etch virus), p19 (from Tomato bushy stunt virus), γb (from Barley stripe mosaic virus) and p21 (from Beet yellows virus) led to an almost twofold increase in initial GFP expression levels, followed by a rapid decline. In contrast, fusions of HCPro, p19, and γb to the 3′-end of GFP resulted in slightly lower but more prolonged GFP expression. Compared with the co-introductions, all GFP::Suppressor translational fusions gave reduced GFP fluorescence levels, suggesting interference of the fusion partner with GFP fluorescence. Regardless of the configuration, introductions of the silencing suppressors AL2 (from Tomato golden mosaic virus) and 126-kDa protein (from Tobacco mosaic virus) resulted in very low GFP fluorescence. This is the first report that directly compares the effects of a large number of viral suppressors of silencing on transient transgene expression using both translational fusions and co-introductions.

Keywords

GFP Lima bean cotyledons Particle bombardment RNA silencing Viral gene-silencing suppressors Gene silencing 

Abbreviations

HCPro

Helper component-proteinase

CaMV

Cauliflower mosaic virus

CMV

Cucumber mosaic virus

TEV

Tobacco etch virus

TBSV

Tomato bushy stunt virus

TGMV

Tomato golden mosaic virus

BYV

Beet yellows virus

BSMV

Barley stripe mosaic virus

TMV

Tobacco mosaic virus

PVX

Potato virus X

Notes

Acknowledgments

The authors acknowledge Dr. Robert A. Bouchard for offering guidance in construct design and providing unique insight on the results of suppressor introductions. The authors also wish to thank Drs. Eric J. Stockinger and David M. Bisaro for their helpful suggestions during the course of this work. Salaries and research support were provided by the United Soybean Board and by State and Federal funds appropriated to The Ohio State University/Ohio Agricultural Research and Development Center. Mention of trademark or proprietary products does not constitute a guarantee or warranty of the product by OSU/OARDC and also does not imply approval to the exclusion of other products that may also be suitable.

Supplementary material

ESM1 (MOV 6216 kb)

ESM2 (MOV 5711 kb)

299_2009_675_MOESM3_ESM.doc (32 kb)
Supplemental Table 1 (DOC 33 kb)
299_2009_675_MOESM4_ESM.doc (31 kb)
Supplemental Table 2 (DOC 31 kb)

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Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Taniya Dhillon
    • 1
  • Joseph M. Chiera
    • 1
  • John A. Lindbo
    • 2
    • 3
  • John J. Finer
    • 1
    Email author
  1. 1.Department of Horticulture and Crop ScienceOARDC/The Ohio State UniversityWoosterUSA
  2. 2.Department of Plant PathologyOARDC/The Ohio State UniversityWoosterUSA
  3. 3.Campbell’s SeedsDavisUSA

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