Abstract
The nucleotide sequence of the pathogenesis-related protein 1(PR-1) gene was obtained from Nicotiana benthamiana using RT-PCR. Restriction enzyme cutting sites of EcoRI and NotI were introduced to the ORF fragments of PR-1, they were then linked together with pET-30a (+) and transformed into E. coli BL21 (DE3). The target protein was induced by 1.5 mM IPTG, at 37°C for 4 h. The expressed protein was purified by Ni–NTA and an anti-NbPR-1 polyclonal antibody was prepared using rabbits. The antibody could detect the expression of PR-1 in N. benthamiana and other Nicotiana plants. NbPR-1 protein has four α-helices and two β-sheets by homology modeling. Furthermore, the purified NbPR-1 protein exhibited a broad-spectrum antifungal activity.
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References
Cutt JR, Harpster MH, Dixon DC, Carr JP, Dunsmuir P, Klessig DF (1989) Disease response to tobacco mosaic virus in transgenic tobacco plants that constitutively express the pathogenesis-related PR1b gene. Virology 173:89–97
Dangl JL, Jones JDG (2001) Plant pathogens and integrated defence responses to infection. Nature 411:826–833
Goodin MM, Zaitlin D, Naidu RA, Lommel SA (2008) Nicotiana benthamiana: its history and future as a model for plant–pathogen interactions. Mol Plant Microbe Interact 21:1015–1026
Kumar D, Klessig DF (2003) High-affinity salicylic acid-binding protein 2 is required for plant innate immunity and has salicylic acid-stimulated lipase activity. Proc Natl Acad Sci USA 100:16101–16106
Li J, Du LF, Liu X, Du SW, Huang XH, Jiang JH (2009) Expression and purification of tobacco PR-1a protein for function analysis. Asian J Chem 21:3697–3707
Loake G, Grant M (2007) Salicylic acid in plant defence–the players and protagonists. Curr Opin Plant Biotechnol 10:466–472
Niderman T, Genetet I, Bruyère T, Gees R, Stintzi A, Legrand M, Fritig B, Mosinger E (1995) Pathogenesis-related PR-1 proteins are antifungal. Plant Physiol 108:17–27
Park SC, Ro Lee J, Kim JY, Hwang I, Nah JW, Cheong H, Park Y, Hahm KS (2010) Pr-1, a novel antifungal protein from pumpkin rinds. Biotechnol Lett 32:125–130
Puthoff DP, Holzer FM, Perring TM, Walling LL (2010) Tomato pathogenesis-related protein genes are expressed in response to Trialeurodes vaporariorum and Bemisia tabaci biotype B feeding. J Chem Ecol 36:1271–1285
Rauscher M, Adam AL, Wirtz S, Guggenheim R, Mendgen K, Deising HB (1999) PR-1 protein inhibits the differentiation of rust infection hyphae in leaves of acquired resistant broad bean. Plant J 19:625–633
Selsa J, Mathysa J, De Conincka BMA, Cammue BPA, De Bolle MFC (2008) Plant pathogenesis–related (PR) proteins: a focus on PR peptides. Plant Physiol Bioch 46:941–950
Seo PJ, Lee AK, Xiang FN, Park CM (2008) Molecular and functional profiling of Arabidopsis pathogenesis-related genes: insights into their roles in salt response of seed germination. Plant Cell Physiol 49:334–344
Tanga W, Zhu SJ, Li LL, Liu DJ, Irving DE (2010) Differential expressions of PR1 and chitinase genes in harvested bananas during ripening, and in response to ethephon, benzothiadizole and methyl jasmonate. Postharvest Biol Tec 57:86–91
van Loon LC, Rep M, Pieterse CMJ (2006) Significance of inducible defense-related proteins in infected plants. Annu Rev Phytopathol 44:135–162
Wang SD, Zhu F, Yuan S, Yang H, Xu F, Shang J, Xu MY, Jia SD, Zhang ZW, Wang JH, Xi DH, Lin HH (2011) The roles of ascorbic acid and glutathione in symptom alleviation to SA-deficient plants infected with RNA viruses. Planta 234:171–181
Xi DH, Yang H, Jiang Y, Xu MY, Shang J, Zhang ZW, Cheng SY, Sang LS, Lin HH (2010) Interference between Tobacco necrosis virus and Turnip crinkle virus in Nicotiana benthamiana. J Phytopathol 158:263–269
Acknowledgments
This work was supported by the National Nature Science Foundation of China (91017004, 30970214, 31070210, 31071669, and 31171835), National Key Basic Research ‘973’ Program of China (2009CB118500), Fundamental Research Funds for the Central Universities (2011SCU04B34) and Doctoral Foundation of the Ministry of Education (20110181110059 and 20070610172).
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Zhu, F., Xu, M., Wang, S. et al. Prokaryotic expression of pathogenesis related protein 1 gene from Nicotiana benthamiana: antifungal activity and preparation of its polyclonal antibody. Biotechnol Lett 34, 919–924 (2012). https://doi.org/10.1007/s10529-012-0851-5
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DOI: https://doi.org/10.1007/s10529-012-0851-5