| Summary: | 碩士 === 國立高雄師範大學 === 生物科學研究所 === 91 === Turnip mosaic virus（TuMV）, a member of Potyvirus genus in Potyviridae family, is one of the most important viruses infecting cruciferous crops, and spreads worldwide including Africa, Asia, Australia, Europe, and North and South America. TuMVs are classified into four strains - C1, C2, C3, and C4 according to various symptoms developed on different Chinese cabbages varieties. A fifth strain, C5, and another special strain, TW, were reported in Taiwan. TuMV-TW and C1 were sequenced in our laboratory in 2001. In this study, modified Moloney Murine Leukemia Virus reverse transcriptase was used to synthesize the full-length first strand cDNAs of TuMV-TW and C1. The 5'' terminal primer was designed to include a T7 promoter sequence and partial viral 5'' NTR sequence. The 3'' terminal primer was designed to include a 14-T tail and a restriction enzyme cutting site. Both primers were used to synthesize full-length double-stranded viral cDNA by PCR. With the completion of the full-length dsDNA, TuMV-TW and C1 transcripts were obtained by in vitro transcription with T7 RNA polymerase. Purificated transcripts were inoculated onto mustard MT1 and confirmed by ELISA 14 and 24 days after inoculation. Mottlings were observed on mustard inoculated leaves with capped TuMV-TW transcripts at day 24. No symptom was observed for TuMV-TW transcripts without cap and TuMV-C1 transcripts with or without cap. ELISA test showed positive reaction on plant 14 and 24 days after inoculation with TuMV-TW transcripts with cap. A 10kb RT-PCR product was confirmed from plants infected with TuMV-TW transcripts with cap.
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