Efficiency of VIGS and gene expression in a novel bipartite potexvirus vector delivery system as a function of strength of TGB1 silencing suppression

• Published in: Virology (New York, N.Y.) Vol. 402; no. 1; pp. 149 - 163
• Main Authors: Lim, Hyoun-Sub, Vaira, Anna Maria, Domier, Leslie L, Lee, Sung Chul, Kim, Hong Gi, Hammond, John
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 20.06.2010
• Subjects: Agroinfiltration; Alternanthera; Alternanthera mosaic virus; Amino Acid Substitution - genetics; Arabidopsis - virology; Arabidopsis thaliana; Bacteriophage T7 - genetics; Bipartite launch system; Cauliflower mosaic virus; Caulimovirus - genetics; coat proteins; Gene Expression; gene expression regulation; Gene Silencing; Genes, Viral; Genetic Vectors; green fluorescent protein; In vivo transcription; Infectious Disease; Mutation, Missense; Nicotiana - virology; Nicotiana benthamiana; Plant viral vector; plant viruses; Potexvirus; Potexvirus - genetics; Potexvirus - immunology; Promoter Regions, Genetic; RNA-Binding Proteins - genetics; RNA-directed DNA polymerase; triple gene block protein; VIGS; viral proteins; virus-induced gene silencing; Plant viral vector; Agroinfiltration; Bipartite launch system; Potexvirus; In vivo transcription; VIGS; Alternanthera mosaic virus
• ISSN: 0042-6822; 1096-0341; 1096-0341
• DOI: 10.1016/j.virol.2010.03.022

We have developed plant virus-based vectors for virus-induced gene silencing (VIGS) and protein expression, based on Alternanthera mosaic virus (AltMV), for infection of a wide range of host plants including Nicotiana benthamiana and Arabidopsis thaliana by either mechanical inoculation of in vitro transcripts or via agroinfiltration. In vivo transcripts produced by co-agroinfiltration of bacteriophage T7 RNA polymerase resulted in T7-driven AltMV infection from a binary vector in the absence of the Cauliflower mosaic virus 35S promoter. An artificial bipartite viral vector delivery system was created by separating the AltMV RNA-dependent RNA polymerase and Triple Gene Block (TGB)123-Coat protein (CP) coding regions into two constructs each bearing the AltMV 5′ and 3′ non-coding regions, which recombined in planta to generate a full-length AltMV genome. Substitution of TGB1 L(88)P, and equivalent changes in other potexvirus TGB1 proteins, affected RNA silencing suppression efficacy and suitability of the vectors from protein expression to VIGS.