| Summary: | 碩士 === 國立臺灣大學 === 植物病理學研究所 === 87 === Calla lilies (Zantedeschia spp.) are perennial bulb plants and belong to the family Araceae. Dasheen mosaic virus (DsMV) and Zantedeschia mosaic virus (ZaMV) which are members of the family Potyviridae, the genus Potyvirus, are frequently found in cultivated calla lilies in Taiwan.
Different calla lily cultivars collected from the field were first checked with ELISA test and EM, and then inoculated to indicator plants. The results indicated that DsMV caused severer symptom than ZaMV did in Philodendron selloum. Besides, different DsMV isolates induced different symptoms in P. selloum. The DsMV-ZAE isolate caused severer symptom than DsMV-TW and DsMV-ZAN did. Chlorotic local lesions appeared on inoculated leaves of Chenopodium quinoa and C. amaranticolor when inoculated with DsMV isolates. DsMV virions from calla lilies were purified and observed with EM. The average length of DsMV-ZAN virions is 754 + 46 nm and that of DsMV-ZAE is 747 + 26 nm. While purified ZaMV-BG virions has the average length of 770 + 75 nm. 1.8 kb genome fragments of DsMV were amplified by RT-PCR. These fragments were cloned and sequenced, which contain a part of the NIb gene, the complete coat protein (CP) gene and the 3’ non-coding region (3’NCR), without poly (A) tail. The sequences of the 3’-terminal 1,824 and 1,825 nucleotides of DsMV-ZAN and DsMV-ZAE were separately determined. The percent similarity of amino acid sequence of the CP gene of DsMV-ZAN and DsMV-ZAE isolates is 99﹪which is higher than other DsMV isolates. According to the sequence analysis, percent similarities of amino acid sequence between the CP gene of DsMV-ZAE and other 10 potyviruses are 59-65%, and percent nucleotide identities of 3’NCR are 20-28%. The 1.6 kb genome fragments of ZaMV was amplified by RT-PCR. According to the sequence analysis, percent similarities of amino acid sequence between the CP gene of ZaMV-BG and other 11 potyviruses are 62-72%, and percent nucleotide identities of 3’NCR are 9-23%.
Among four sample extraction buffers, GE buffer was the best one when using DAS-ELISA to detect DsMV. Symptom severities were related to the concentrations of DsMV in different leaf tissues of P. selloum. Two specific probes were designed which located in 3’NCR of DsMV and ZaMV due to the low homology. The sensitivities of four specific probes were about 8 pg for the plasmid DNA of DsMV and ZaMV clones. The sensitivity of DsMV-specific probe, DCP+3’NCR, was up to 4 ng total RNA of DsMV-ZAE; and only 20 ng for D3’NCR probe when detected with dot-blot hybridization. The sensitivity of ZaMV-specific probe, ZCP, was about 20 ng total RNA. When compared the sensitivity of DAS-ELISA, DsMV-specific probes and RT-PCR for detecting purified DsMV-ZAN, the results indicated that hemi-nested RT-PCR was about 96,450 times more sensitive than ELISA, and specific probes was about 3 — 77 times more sensitive than ELISA.
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