芜菁黄化花叶病毒RT-LAMP检测方法的建立
Establishment of the Detection Method for Turnip yellow mosaic virus by RT-LAMP
DOI: 10.12677/BP.2022.122013, PDF,   
作者: 杨馥韩, 雷云廷, 杨再福, 冯文卓, 王 勇, 陈相儒*：贵州大学农学院，贵州 贵阳；姜 宁：云南烟草科学研究院，云南 昆明
关键词: 芜菁黄化花叶病毒；RT-LAMP；病毒检测； Turnip yellow mosaic virus (TYMV)； RT-LAMP； Virus Detection
摘要: 芜菁黄化花叶病毒(Turnip yellow mosaic virus, TYMV)是芜菁黄花叶病毒科(Tymoviridae)芜菁黄花叶病毒属(Tymovirus)的代表种，该病毒侵染白菜能够引起黄化、花叶和皱缩等症状，给白菜产业造成巨大的影响。在此，我们建立了反转录环介导等温扩增(RT-LAMP)技术来检测白菜中的TYMV。针对TYMV分离株中保守的RNA依赖性RNA聚合酶(RdRp)基因的区域设计的RT-LAMP检测引物。优化后的RT-LAMP方法对TYMV的检测具有特异性和灵敏度，灵敏度比RT-PCR高10,000倍。这种简单、高效、快速和灵敏的方法能够应用于白菜中TYMV的检测。
Abstract: Turnip yellow mosaic virus (TYMV) is the type species of the genus Tymovirus in Tymoviridae. TYMV induce symptoms such as yellowing, mosaic and leaf rugosity in Chinese cabbage, damaging the Chinese cabbage industry. Therefore, developing an easy, efficient strategy for detecting TYMV is crucial. Here, we established a two-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) technique for detecting TYMV in Chinese cabbage. RT-LAMP primers were designed against regions of the RNA-dependent RNA polymerase (RdRp) gene conserved among TYMV isolates. The optimized RT-LAMP method showed high specificity and sensitivity to TYMV, displaying 10,000 times greater sensitivity than conventional RT-PCR. This easy, efficient, sensitive, and inexpensive method for detecting TYMV in Chinese cabbage is the first application of RT-LAMP, to our knowledge, for detecting TYMV.
文章引用：杨馥韩, 雷云廷, 姜宁, 杨再福, 冯文卓, 王勇, 陈相儒. 芜菁黄化花叶病毒RT-LAMP检测方法的建立[J]. 生物过程, 2022, 12(2): 116-123. https://doi.org/10.12677/BP.2022.122013

参考文献

[1] Mossop, D.W., Fry, P.R. and Young, B.R. (1983) New Plant Disease Records in New Zealand: Arabis Mosaic Virus in Celery, Lettuce and Chinese Cabbage, Tomato Spotted Wilt Virus in Celery. New Zealand Journal of Agricultural Re-search, 26, 257-259. [Google Scholar] [CrossRef] 
[2] Cai, L., Zhang, Y.C., Chen, Y.S., Hong, N. and Xu, W.X. (2017) First Report of Broad Bean Wilt Virus 2 Infecting Non-Heading Chinese Cabbage in China. Plant Disease, 101, 1559-1560. [Google Scholar] [CrossRef] 
[3] Farzadfar, S.H., Ahoomanesh, A., Mosahebi, G.H., Pour-rahim, A.R. and Golnaraghi, A.R. (2007) Occurrence and Distribution of Cauliflower Mosaic Virus on Cruciferous Plants in Iran. Plant Pathology Journal, 6, 22-29. [Google Scholar] [CrossRef] 
[4] Liu, S., Cao, X. and Yuan, X. (2017) First Report of Cucumber Mo-saic Virus and Its Associated Satellite RNA in Celery Cabbage in Shandong Province of China. Plant Disease, 101, 1829-1830. [Google Scholar] [CrossRef] 
[5] Kim, J.S., Cho, J.D., Choi, H.S., Lee, S.H., Choi, G.S., Lee, S.Y., Kim, H.J. and Yoon, M.K. (2010) Ribgrass Mosaic Tobamovirus Occurred on Chinese Cabbage in Korea. The Plant Pathology Journal, 26, 328-339. [Google Scholar] [CrossRef] 
[6] Green, S.K. and Deng, T.C. (1985) Turnip Mosaic Virus Strains in Cruciferous Hosts in Taiwan. Plant Disease, 69, 28-31. [Google Scholar] [CrossRef] 
[7] Chen, Y.K. and Chang, Y.S. (2011) First Report of Turnip Ringspot Virus in Field Mustard (Brassica chinensis) in Taiwan. Plant Dis-ease, 95, 1036. [Google Scholar] [CrossRef] 
[8] Kirino, N., Inoue, K., Tanina, K., Yamazaki, Y. and Ohki, S.T. (2008) Turnip yellow mosaic virus Isolated from Chinese Cabbage in Japan. Journal of General Plant Pa-thology, 74, Article No. 331. [Google Scholar] [CrossRef] 
[9] Xiang, H.Y., Dong, S.W., Shang, Q.X., Zhou, C.J., Li, D.W., Yu, J.L. and Han, C.G. (2011) Molecular Characterization of Two Genotypes of a New Polerovirus Infecting Brassicas in China. Archives of Virology, 156, 2251-2255. [Google Scholar] [CrossRef] [PubMed]
[10] Morch, M.D., Boyer, J.C. and Haenni, A.L. (1988) Overlapping Open Reading Frames Revealed by Complete Nucleotide Sequencing of Turnip yellow mosaic virus Genomic RNA. Nu-cleic Acids Research, 16, 6157-6173. [Google Scholar] [CrossRef] [PubMed]
[11] Alfaro-Fernandez, A., Serrano, A., Tornos, T., Cebrian, M.C., Cordo-ba-Selles, M.C., Jorda, C. and Font, M.I. (2016) Turnip yellow mosaic virus in Chinese Cabbage in Spain: Commercial Seed Transmission and Molecular Characterization. European Journal of Plant Pathology, 146, 433-442. [Google Scholar] [CrossRef] 
[12] Spak, J., Kubelkova, D.A. and Hnilicka, E. (1993) Seed Trans-mission of Turnip yellow mosaic virus in Winter Turnip and Winter Oil Seed Rapes. Annals of Applied Biology, 123, 33-35. [Google Scholar] [CrossRef] 
[13] de Assis Filho, F.M. and Sherwood, J.L. (2000) Evalua-tion of Seed Transmission of Turnip yellow mosaic virus and Tobacco Mosaic virus in Arabidopsis thaliana. Phyto-pathology, 90, 1233-1238. [Google Scholar] [CrossRef] 
[14] Fukuta, S., Ohishi, K., Yoshida, K., Mizukami, Y., Ishida, A. and Kanbe, M. (2004) Development of Immunocapture Reverse Transcription Loop-Mediated Isothermal Amplifica-tion for the Detection of Tomato Spotted Wilt Virus from Chrysanthemum. Journal of Virological Methods, 121, 49-55. [Google Scholar] [CrossRef] [PubMed]
[15] Buates, S., Bantuchai, S., Sattabongkot, J., Han, E.T., Tsuboi, T., Udomsangpetch, R., Sirichaisinthop, J. and TanAriya, P. (2010) Development of a Reverse Transcrip-tion-Loop-Mediated Isothermal Amplification (RT-LAMP) for Clinical Detection of Plasmodium Falciparum gameto-cytes. Parasitology International, 59, 414-420. [Google Scholar] [CrossRef] [PubMed]
[16] Fukuta, S., Iida, T., Mizukami, Y., Ishida, A., Ueda, J., Kanbe, M. and Ishimoto, Y. (2003) Detection of Japanese Yam Mosaic Virus by RT-LAMP. Archives of Virology, 148, 1713-1720. [Google Scholar] [CrossRef] [PubMed]
[17] Varga, A. and James, D. (2006) Use of Reverse Transcription Loop-mediated Isothermal Amplification for the Detection of Plum Pox Virus. Journal of Virological Methods, 138, 184-190. [Google Scholar] [CrossRef] [PubMed]
[18] Peng, Q., Ning, J., Xu, Q., Yang, T., Wang, Y., Zheng, T., Zhuang, Q. and Xi, D. (2021) Development and Application of a Reverse Transcription Loop-Mediated Iso-thermal Amplification Combined with Lateral Flow Dipstick for Rapid and Visual Detection of Citrus Leaf Blotch Virus in Kiwifruit. Crop Protection, 143, Article ID: 105555. [Google Scholar] [CrossRef] 
[19] Feng W Z, Hieno, A., Kusunoki, M., Suga, H. and Kageyama, K. (2018) LAMP Detection of Four Plant-Pathogenic Oomycetes and Its Application in Lettuce Fields. Plant Disease, 103, 298-307. [Google Scholar] [CrossRef] 
[20] Wang, J., Hou, X., Yang, X. and Gustafson, P. (2011) Identi-fication of Conserved MicroRNAs and Their Targets in Chinese Cabbage (Brassica rapa Subsp. Pekinensis). Genome, 54, 1029-1040. [Google Scholar] [CrossRef] [PubMed]
[21] Liu, Y., Li, F., Li, Y.Y., Zhang, S.B., Gao, X.W., Xie, Y., Yan, F., Zhang, A.S., Dai, L.Y., Cheng, Z.B., Ding, M., Niu, Y.B., Wang, S.B., Che, H.Y., Jiang, T., Shi, X.B., He, Z.F., Wu, Y.F., Zhang, D.Y., Qing, L., Yan, W.R., Yang, X.H., Tang, Y.F., Zheng, H.Y., Tang, Q.J., Zhang, S.B., Zhang, D.F., Cai, L. and Tao, X.R. (2019) Species, Distribution and Occurrence Trends of Viruses Infesting Major Veg-etable Crops in China. Chinese Agricultural Science, 52, 239-261.
[22] Kokane, A.D., Kokane, S.B., Warghane, A.J., Gubyad, M.G., Sharma, A.K., Reddy, M.K. and Ghosh, D.K. (2021) A Rapid and Sensitive Reverse Transcrip-tion-Loop-Mediated Isothermal Amplification (RT-LAMP) Assay for the Detection of Indian Citrus Ringspot Virus. Plant Disease, 105, 1346-1355. [Google Scholar] [CrossRef]