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Abstract Tomato mosaic virus (TAV) is an important pathogen in agriculture, infecting more than 100 species of 24 dicotyledoneae and 3 monocotyledoneae, such as Chenopodiaceae and Solanaceae. In order to analyze the role of genomic RNAs, full length sequence of TAV isolated from Beijing (TAV-BJ) was obtained by RT-PCR and infectious cDNA clones was constructed. Using total RNA from Nicotiana glutinosa infected by TAV-BJ as template, full length of RNA2 and RNA3 were amplified by RT-PCR, and RNA1 was obtained from cDNA of TAV-BJ double stand RNA1. PCR products of RNA1, RNA2 and RNA3 were cloned into pUC118 vector and sequenced. TAV-BJ RNA1 contained 3 409 nucleotides (nts), encoding 1a protein of 994 amino acids, RNA2 was 3 032 nts, encoding 2a protein of 829 amino acids and 2b protein of 78 amino acids and RNA3 consisted of 2 218 nts and encoded 3a protein of 247 amino acids and coat protein (CP) of 219 amino acids (GenBank accession No of TAV-BJ genomic RNA1, 2 and 3 was HQ424163, HQ424164 and HQ424165, respectilvely). cDNA clones of TAV-BJ genomic RNAs were transcribed into RNA and the mixture of transcripts RNA was mechanically inoculated onto N. glutinosa. At days post-inoculation (dpi), T1T2T3 induced host plants to express mosaic symptom, which was in accordance with that appearing on N. glutinosa infected by TAV-BJ viral RNAs. Seedlings of N. glutinosa infected by CMV-Fny lack of 2b gene (CMV-FnyΔ2b) was symptomless. Co-inoculation of T1, T2, T2Δ2b or T3 respectively with CMV-FnyΔ2b showed that the pseudorecombinant virus with T2 or T3 could resue the pathogenicity of CMV-FnyΔ2b in host plant, thus RNA3 of TAV-BJ might paly some function of 2b protein of CMV-FnyΔ2b. TAV-BJ RNA3 chimera infectious clones that replaced with 3a or CP gene fragment of CMV-Fny were constructed by Overlapping PCR. Biological assays of F1F2Δ2bRNA3T3aFcp and F1F2Δ2bRNA3 F3aTcp suggested that TAV 3a protein could compensate some function of 2b protein.In this study, infectious cDNA clones of TAV-BJ are successfully constructed and results of pseudorecombinant virus between CMV-Fny and TAV-BJ confirm that 3a gene of TAV-BJ have some role of CMV-Fny 2b gene.
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Received: 16 May 2011
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