Abstract
We have utilized plant-based transient expression to produce tobacco mosaic virus (TMV)-based nano-rods of predetermined lengths. This is achieved by expressing RNAs containing the TMV origin of assembly sequence (OAS) and the sequence of the TMV coat protein either on the same RNA molecule or on two separate constructs. We show that the length of the resulting nano-rods is dependent upon the length of the RNA that possesses the OAS element. By expressing a version of the TMV coat protein that incorporates a metal-binding peptide at its C-terminus in the presence of RNA containing the OAS we have been able to produce nano-rods of predetermined length that are coated with cobalt-platinum. These nano-rods have the properties of defined-length nano-wires that make them ideal for many developing bionanotechnological processes.
Highlights
Particles of Tobacco mosaic virus (TMV) consist of a 6.4 kb molecule of single-stranded RNA encapsidated by approximately 2130 copies of the 17.5 kDa coat protein arranged with helical symmetry
Using the pEAQ-HT vector system (Sainsbury et al, 2009) to express both the tobacco mosaic virus (TMV) coat protein and RNA molecules with the OAS positioned near the 3 terminus, we demonstrate the efficient production of rods of predetermined length and, by the expression of variant coat protein, the generation of nanorods that promote the binding of cobalt-platinum to the exterior surface, thereby producing conducting nano-wires
As a first step to creating defined length nano-rods, we designed a DNA sequence that included the TMV coat protein open reading frame with a TMV OAS element to its 3 side. This reverses the order of the OAS and coat protein (CP) that is found in TMV U1 genome and was done to ensure that the majority of RNA encapsidation would occur in the fast 3 – 5 direction
Summary
Particles of Tobacco mosaic virus (TMV) consist of a 6.4 kb molecule of single-stranded RNA encapsidated by approximately 2130 copies of the 17.5 kDa coat protein arranged with helical symmetry. NuPAGE gel electrophoresis and Instant Blue staining revealed the presence of a single band of the size expected for TMV coat protein in the nano-rod-containing fractions of the CsCl gradients in the case of pEAQ-HT-TMV-CP/OAS and pEAQHT-TMV-CP/VP60/OAS (Figure 2B, lanes 1 and 2).
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