Abstract
Transgenic rapeseed (Brassica napus L.) plants carrying an artificial gene for the antimicrobial peptide cecropin P1 (cecP1) were obtained and characterized. The agrobacterial transformation was done by vacuum infiltration of seeds with agrobacterium GV3101(pMP90RK) containing a binary vector pGA482::cecP1. The cec1 gene expression was analyzed by Western blotting and confirmed by antimicrobial activity measurements of plant extracts. The obtained plants showed the resistance to the bacterial and fungal pathogens Erwinia carotovora and Fusarium sporotrichioides. The photosynthetic activities of control and transgenic plants under biotic stress conditions of E. carotovora infection were comparatively studied. The higher tolerance of the cecP1 plants to the oxidative stress caused by paraquat was shown. The results obtained point to the possibility of incorporation of the cecropin P1 gene into the integral stress protection system of plants.
Similar content being viewed by others
Abbreviations
- cecP1:
-
cecropin P1
- DETC:
-
diethylthiocarbamate
- DF:
-
delayed fluorescence of chlorophyll a
- Km:
-
kanamycin
- NBT:
-
nitoblue tetrazolium
- nptII :
-
neomycintransferase II
- SOD:
-
superoxide dismutase
References
Chavadej, S., Brisson, N., McNeil, J.N., and Luca, V., Redirection of Tryptophan Leads to Production of Low Indole Glucosinolate Canola, Proc. Natl. Acad. Sci. USA, 1994, vol. 91, pp. 2166–2170.
Falco, S.C., Guida, T., Locke, M., Mauvais, T., Sanders, C., Ward, R.T., and Webber, P., Transgenic Canola and Soybean Seeds with Increased Lysine, Biotechnology, 1995, vol. 13, pp. 577–582.
Grison, R., Grezes-Besset, B., Schneider, M., Lucante, N., Olsen, L., Leguay, J.J., and Toppan, A., Field Tolerance to Fungal Pathogens of Brassica napus Constitutively Expressing a Chimeric Chitinase Gene, Nat. Biotechnol., 1996, vol. 14, pp. 643–646.
Carpenter, J. and Gianessi, L., Herbicide Use on Roundup Ready Crops, Science, 2000, vol. 287, pp. 803–804.
Parashina, E.V., Serdobinskii, L.A., Kalle, E.G., Lavrova, N.V., Avetisov, V.A., Lunin, V.G., and Naroditskii, B.S., Genetic Engineering of Oilseed Rape and Tomato Plants Expressing a Radish Defensin Gene, Russ. J. Plant Physiol., 2000, vol. 47, pp. 417–423.
Zakharchenko, N.S., Rukavtsova, E.B., Gudkov, A.T., and Buryanov, Ya.I., Enhanced Resistance to Phytopathogenic Bacteria in Transgenic Tobacco Plants with Synthetic Gene of Antimicrobial Peptide Cecropin P1, Russ. J. Genetics, 2005, vol. 41, pp. 1187–1193.
Murashige, T. and Skoog, F., A Revised Medium for Rapid Growth and Bioassays with Tobacco Cultures, Physiol. Plant., 1962, vol. 15, pp. 473–497.
An, G., Ebert, P., Mitra, A., and Ha, S.B., Binary Vectors, Plant Molecular Biology Manual, Gelvin, S.B., Schilperoort, R.A., and Verma, D.P, Eds., Dordrecht: Kluwer, 1988, Ch. A3, pp. 1–19.
Martemyanov, K.A., Shirokov, V.A., Kurnasov, O.V., Gudkov, A.T., and Spirin, A.S., Cell-Free Production of Biologically Active Polypeptides: Application to the Synthesis of Antibacterial Peptide Cecropin, Protein Expres. Pur., 2001, vol. 21, pp. 456–461.
Maniatis, T., Frisch, E.F., and Sambrook, J., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor: Cold Spring Harbor Lab., 1982.
Bechtold, D., Ellis, J., and Pelletier, G., In Planta Agrobacterium Mediated Gene Transfer by Infiltration of Adult Arabidopsis thaliana Plants, C.R. Acad. Sci., Life Sci., 1993, vol. 316, pp. 1194–1199.
Edwards, K., Johnstone, C., and Thompson, C., A Simple and Rapid Method for the Preparation of Plant Genomic DNA for PCR Analysis, Nucleic Acids Res., 1991, vol. 19, pp. 1349–1351.
Jan, P.S., Huang, H.Y., and Chen, H.M., Expression of a Synthesized Gene Encoding Cationic Peptide Cecropin B in Transgenic Tomato Plants Protects against Bacterial Diseases, Appl. Environ. Microbiol., 2010, vol. 2, pp. 769–775.
Schagger, H. and von Jagow, G., Tricine-Sodium Dodecyl Syfate-Polyacrylamide Gel Electrophoresis for the Separation of Proteins in the Range from 1 to 100 kDa, Anal. Biochem., 1987, vol. 166, pp. 368–379.
Ohshima, M., Mitruhara, I., Okamoto, M., Sawano, S., Nishiyama, K., Kaku, H., Natori, S., and Ohashi, Y., Enhanced Resistance to Bacterial Diseases of Transgenic Tobacco Plants Overexpressing Sarcotoxin IA, a Bactericidal Peptide of Insect, J. Biochem., 1999, vol. 125, pp. 431–435.
Draper, J., Scott, R., and Hamil, J., Plant Genetic Transformation and Gene Expression, A Laboratory Manual, Draper, J., Scott, R., and Armitage, P., Walden, R., Eds., Oxford: Blackwell Sci. Publ., 1988, pp. 69–160.
Priol, J.L. and Chartier, P., Partitioning of Transfer and Carboxylation Components of Intracellular Resistance to Photosynthetic CO2 Fixation: A Critical Analysis of the Methods Used, Ann. Bot., 1977, vol. 41, pp. 789–800.
Lavorel, J., Fast and Slow Phases of Luminescence in Chlorella, Photochem. Photobiol., 1975, vol. 21, pp. 331–343.
Bigler, W. and Schreiber, U., Chlorophyll Luminescence as an Indicator of Stress-Induced Damage to the Photosynthetic Apparatus. Effects of Heat-Stress in Isolated Chloroplasts, Photosynth. Res., 1990, vol. 25, pp. 161–171.
Veselovskii, V.A. and Veselova, T.V., Lyuminestsentsiya rastenii (Plant Luminescence), Moscow: Nauka, 1990.
Goltsev, V., Chernev, P., Zaharieva, I., Lambrev, P., and Strasser, R.J., Kinetics of Delayed Chlorophyll a Fluorescence Registered in Milliseconds Time Range, Photosynth. Res., 2005, vol. 84, pp. 209–215.
Bellincampi, D., Dipierro, N., Salvi, G., Cervone, F., and de Lorenzo, G., Extracellular H2O2 Induced by Oligogalacturonides Is Not Involved in the Inhibition of the Auxin-Regulated rolB Gene Expression in Tobacco Leaf Explants, Plant Physiol., 2000, vol. 122, pp. C. 1379–1385.
Chaitanya, K.S.K., Naithani, S.C., and Faertn, F., Role of Superoxide Lipid Peroxidation and Superoxide Dismutation in Membrane Perturbation during Loss of Viability in Seeds of Shorea robusta, New Phytol., 1994, vol. 126, pp. 623–627.
Beauchamp, C.O. and Fridovich, I., Superoxide Dismutase: Improved Assays and an Assay Applicable to Acrylamide Gels, Ana. Biochem., 1971, vol. 44, pp. 276–287.
Wraight, C.A. and Crofts, A.R., Delayed Fluorescence and Light-Energy State of Chloroplasts, Eur. J. Biochem., 1971, vol. 19, pp. 386–397.
Zakharchenko, N.S., Rukavtsova, E.B., Gudkov, A.T., Yukhmanova, A.A., Shkolnaya, L.A., Kado, K.I., and Buryanov, Ya.I., Expression of the Artificial Gene Encoding Anti-Microbial Peptide Cecropin P1 Increases the Resistance of Transgenic Potato Plants to Potato Blight and White Rot, Dokl. Biol. Sci., 2007, vol. 415, pp. 267–269.
Zakharchenko, N.S., Lebedeva, A.A., and Buryanov, Ya.I., Technique for Production of Transgenic Kalanchoe Plants Expressed Cecropin P1 Gene, RF Patent no. 2445768, Byull. Izobret., 2012, no. 9.
Mittova, V., Tal, M., Volokita, M., and Guy, M., UpRegulation of the Leaf Mitochondrial and Peroxisomal Antioxidative Systems in Response to Salt-Induced Oxidative Stress in the Wild Salt-Tolerant Tomato Species Lycopersicon pennellii, Plant Cell Environ., 2003, vol. 26, pp. 845–856.
Kreslavskii, V.D., Los, D.A., Allakhverdiev, S.I., and Kuznetsov, Vl.V., Signaling Role of Reactive Oxygen Species in Plants under Stress, Russ. J. Plant Physiol., 2012, vol. 59, pp. 141–154.
Campo, S., Manrique, S., Garcia-Martinez, J., and San Segundo, S., Production of Cecropin A in Transgenic Rice Plants Has an Impact on Host Gene Expression, Plant Biotechnol. J., 2008, vol. 6, pp. 585–608.
Author information
Authors and Affiliations
Corresponding author
Additional information
Original Russian Text © N.S. Zakharchenko, Ya.I. Buryanov, A.A. Lebedeva, S.V. Pigoleva, D.V. Vetoshkina, E.V. Loktyushov, M.A. Chepurnova, V.D. Kreslavski, A.A. Kosobryukhov, 2013, published in Fiziologiya Rastenii, 2013, Vol. 60, No. 3, pp. 424–433.
Rights and permissions
About this article
Cite this article
Zakharchenko, N.S., Buryanov, Y.I., Lebedeva, A.A. et al. Physiological features of rapeseed plants expressing the gene for an antimicrobial peptide cecropin P1. Russ J Plant Physiol 60, 411–419 (2013). https://doi.org/10.1134/S1021443713030163
Received:
Published:
Issue Date:
DOI: https://doi.org/10.1134/S1021443713030163