Segmented GFP-like aptamer probes for functional imaging of viral genome trafficking.
Recently developed GFP-like RNA aptamers harbor a few unique potential benefits for in vivo RNA imaging applications, including co-packaging of viral genomes. Here we examine them in the context of co-packaging of RNA strands during virion assembly and trafficking. The approach is applicable both in vitro and in vivo, thus bridging an existing methodological gap. We have found that splitting the aptamer sequence in the loop region into two separate parts allows for subsequent self-assembly into a functional unit, which preserves the dye-binding pocket. In presence of the dye, virus-like particles encapsulating segmented GFP-like aptamers provided bright fluorescence emission and showed negligible bleaching due to continuous chromophore exchange: two desirable characteristics for real-time in vivo single particle studies requiring a broader dynamic range than currently available. Proof-of-principle in vivo imaging experiments confirmed detectability of aptamer-loaded virus-like particles in barley root cells even in presence of significant autofluorescence background.
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA. Electronic address: itsvetko@indiana.edu.
Irina B
IB
Tsvetkova
Molecular and Cellular Biochemistry Department, Indiana University, Bloomington, Indiana 47405, USA.
Guanghui
G
Yi
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA.
Yi
Y
Yi
Molecular and Cellular Biochemistry Department, Indiana University, Bloomington, Indiana 47405, USA.
Cheng C
CC
Kao
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA. Electronic address: dragnea@indiana.edu.
Bogdan G
BG
Dragnea
D052157:Aptamers, Nucleotide / Q000378:metabolism*; D017795:Bromovirus / Q000235:genetics / Q000502:physiology*; D001467:Hordeum / Q000821:virology; D061848:Optical Imaging / Q000379:methods*; D018517:Plant Roots / Q000821:virology; D012367:RNA, Viral / Q000032:analysis*; D013194:Staining and Labeling / Q000379:methods*; D019065:Virus Assembly
D052157:Aptamers, Nucleotide; D012367:RNA, Viral
Brome mosaic virus; GFP-like RNA aptamer; Viral genome packaging; Virus tracking