Hindawi Publishing Corporation Journal of Biomedicine and Biotechnology Volume 2009, Article ID 781712, 8 pages doi:10.1155/2009/781712 Research Article Virus-Specific Read-Through Codon Preference Affects Infectivity of Chimeric Cucumber Green Mottle Mosaic Viruses Displaying a Dengue Virus Epitope Pak-Guan Teoh, 1 Aik-Seng Ooi, 1 Sazaly AbuBakar, 2 and Rofina Yasmin Othman 1 1 Center for Research in Biotechnology for Agriculture (CEBAR) and Institute of Biological Sciences, Faculty of Science, University of Malaya, Kuala Lumpur 50603, Malaysia 2 Department of Medical Microbiology, Faculty of Medicine, University of Malaya, Kuala Lumpur 50603, Malaysia Correspondence should be addressed to Rofina Yasmin Othman, yasmin@um.edu.my Received 12 August 2008; Accepted 7 January 2009 Recommended by George E. Plopper A Cucumber green mottle mosaic virus (CGMMV) was used to present a truncated dengue virus type 2 envelope (E) protein binding region from amino acids 379 to 423 (EB4). The EB4 gene was inserted at the terminal end of the CGMMV coat protein (CP) open reading frame (ORF). Read-through sequences of TMV or CGMMV, CAA-UAG-CAA-UUA, or AAA-UAG-CAA-UUA were, respectively, inserted in between the CP and the EB4 genes. The chimeric clones, pRT, pRG, and pCG+FSRTRE, were transcribed into full-length capped recombinant CGMMV transcripts. Only constructs with the wild-type CGMMV read-through sequence yielded infectious viruses following infection of host plant, muskmelon (Cucumis melo) leaves. The ratio of modified to unmodified CP for the read-through expression clone developed was also found to be approximately 1:1, higher than what has been previously reported. It was also observed that infectivity was not affected by differences in pI between the chimera and its wild counterpart. Analysis of recombinant viruses after 21-days-postinculation (dpi) revealed that deletions occurred resulting in partial reversions of the viral population to near wild type and suggesting that this would be the limiting harvest period for obtaining true to type recombinants with this construct. Copyright © 2009 Pak-Guan Teoh et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 1. Introduction The development of plant virus vectors as in planta expres- sion systems for foreign genes provides an attractive alter- native biotechnological approach for peptide expression [1– 5]. This method has been exploited in vaccine production, where small foreign peptides are expressed as a fusion with the viral coat proteins. Essentially, an insertion site has to be determined in the virus genome so that the resulting product will be displayed on the surface of the virus particle which is then propagated in plants and consequently isolated and used as antigen presenting vehicles [5, 6]. Modifications that do not interfere with the normal functions of the particular virus are a prerequisite for this peptide fusion approach. One strategy suggests that foreign gene segments could be fused to the terminus of a viral gene in a way that permits the produc- tion of both the fusion product and the native viral protein, thus avoiding interference with normal gene functions. The success of this epitope presentation strategy depends on a detailed knowledge of virus structure at the atomic level, which is only available for a limited number of viruses. We have recently developed Cucumber green mottle mosaic virus (CGMMV) as a candidate for expressing anti- genic peptides in plants [7]. CGMMV is a tobamovirus with a genome size of ∼6.4 kb which has been well characterized both biologically [8, 9] and structurally [10, 11]. In this study, a truncated dengue virus type 2 envelope (E) protein binding region from amino acid 379 to 423 (EB4) was inserted into the end of the coat protein (CP) open reading frame (ORF) of a previously constructed CGMMV full- length clone, pCGT7X [7]. The antigenic peptide was chosen based on a recent study that suggests its importance in enabling dengue virus to bind to specific host cell receptors (S. Abu Bakar personal communication). The present study explores the possibility of extrapolating the CGMMV anti- genic epitope presentation system for developing diagnostics