The Tobacco mosaic virus (TMV) is a positive sense single stranded RNA virus and is found across the world. TMV can impact the overall yield and quality of the crop resulting in an economic loss. Plants that are infected with TMV show a variety of symptoms such as mosaic pattern, mottling, necrotic lesions and stunted growth. Historically, TMV has caused controversy on whether this economically significant virus is seedborne or seed transmitted. The objective of this study is to track TMV infection from infected seeds to seedlings to determine the percentage of seed transmission. This experiment used three pods from three different TMV infected cultivar K 326 flue-cured tobacco plants. Seeds from each pod were germinated in a growth chamber for approximately ten days. Samples were separated into seed coat, root and leaves after germination. Total RNA was extracted from each part and synthesized into cDNA for analysis. A quantitative real-time PCR (RT-qPCR) assay was used to determine TMV concentration of each sample. Endpoint RT-PCR was used to determine a conservative threshold value from the RT-qPCR results. These results demonstrated that TMV influenced percent germination with a range from 94% to 50%. Seed coats had a significantly higher virus titer concentration (P < 0.05) when compared to the roots and leaves. Statistical analysis revealed highly significant (P < 0.0001) differences among pods for virus titer and there is a highly significant plant by pod interaction (P < 0.0001). Endpoint RT-PCR confirmed TMV infection in leaves, roots and seed coats. Percent infection in leaves ranged from 2% to 24% and percent infection for roots ranged from 8% to 40%. Results demonstrate that TMV is seed transmitted in flue-cured tobacco. / Master of Science / The Tobacco mosaic virus (TMV) is an RNA virus that occurs globally in areas where tobacco is grown. TMV is a tobamovirus and infects over 350 plant species. TMV can reduce the yield and quality of the crop which will result in an economic loss for the grower. Plants that are infected with TMV show a variety of symptoms such as mosaic pattern, necrotic lesions, and stunted growth, and there are no effective ways to eradicate the virus. There has been controversy on whether to categorize TMV as a seedborne virus or a seed-transmitted virus because the location of the virus within a seed is unknown. This study examined seeds from three pods grown on three different TMV-infected flue-cured tobacco plants of cultivar K 326 to track TMV infection from infected seeds to seedlings. Seeds from each pod were germinated in a growth chamber for ten days and samples were separated into leaves, root and seed coat. Each sample had total RNA extracted and synthesized into cDNA for analysis. A quantitative real-time PCR (RT-qPCR) assay was used to determine TMV concentration of each sample since this technology can detect small amounts of virus. Endpoint RT-PCR was used to conservatively determine an infection threshold value from the RT-qPCR results. Percent germination of TMV infected seeds ranged from 94% to 50%. Seed coats had a significantly higher virus titer (P < 0.05) when compared to the roots and leaves in each pod. Statistical analysis showed (P < 0.0001) differences among pods for virus titer and there is a highly significant plant by pod interaction (P < 0.0001). Endpoint RT-PCR confirmed TMV infection in leaves, roots and seed coats. Percent infection in iv leaves ranged from 2% to 24% and percent infection for roots ranged from 8% to 40%. Therefore, results show that TMV is seed-transmitted in flue-cured tobacco.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/101702 |
| Date | 10 July 2019 |
| Creators | Hoak, Jessica |
| Contributors | Crop and Soil Environmental Sciences, Wilkinson, Carol A., Sit, Tim Lawrence, Reed, T. David, Welbaum, Gregory E. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Thesis |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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