Rapid method with high efficiency to transform alfalfa for expression of BANYULS (BAN) gene

Document Type : Research Paper

Authors

1 Research Institute of Forests and Rangelands, P.O. Box 13185-116, Tehran, Iran.

2 Tehran University

3 Genetic and Biotechnology Research Institiue of Forage crops, Italy

Abstract

Condensed tannins (CTS) are flavonoid oligomers, many of which have beneficial effects on animal (safe bloat) and human health. The functions of CT are due to their ability to form reversible complex with proteins, the formation of these protein-tannin complexes making protein unavailable for microbial enzyme and activate other mechanisms like deprivation of metal ions and inhibition of oxidative phosphorylation which also results in the prevention of foam formation and bloating. Anthocyanins are flavonoids that contribute to the red, violet or blue pigmentation of both seeds and flowers. The BAN gene encodes anthocyanidin reductase (ANR), an enzyme proposed to convert anthocyanidins to their corresponding 2, 3-cis -flavan -3-ols ((+)-catechin). Transformation of leaf explants from Medicago sativa var. Rgsy27 and M.sativa var. P1 genotypes was carried out with the plasmid pBI121.BAN(binary vector containing the BAN gene) by two strains of Agrobacterium tumefaciens(LBA 4404 & EHA 105). Leaf explants of Rgsy27 produced more than 90% proembryogenic calli, but in P1 produced about 30% .More than 65% of regenerated embryos were changed to whole plants in both genotypes. The efficiency of transformation in Rgsy27 and P1 plants was depended on Agrobacterium strains, so that the strain LBA 4404 was efficient more than EHA 105 for transformation of both genotypes. Ectopic expression of the BAN in Alfalfa transgenic foliage resulted in accumulation of CTs. Thus, it has been assumed that the BAN gene also acts as starter units in tannins condensation in alfalfa. Loss of  the BAN gene function increases the amount of red pigment, suggesting that the BANYULS protein might be a negative regulator of pigment production. BANYULS activity is limiting factor for CT production, making this enzyme an attractive target for induction of over expression in forage plants, such as alfalfa (Medicago sativa) with the goal of inducing CT production in their foliage. The condensed tannin percentage in dry matter of transgenic Rgsy27 and P1 plants was 0.474 and 0.600, respectively and 0.259 and 0.408 in control plants.

Keywords

Main Subjects

References

حسام زاده حجازی ،س.م. و همکاران 1384.روش تراریزش سریع با کارایی بالا در یونجه های دیپلوئید و تتراپلوئید برای تظاهر ژن Sn و بتا-گلوکورونیداز(gus). مجله علوم کشاورزی ایران، جلد36، شماره4.
Blondon, F., Marie , D.,  Brown, S., Kondorosi, A., 1994. Genome Size and base Composition in Medicago sativa and M.truncatula species. Genome, 37: 264-270.
Bloor, S.J, Bradley, J.M, Lewis, D.H, Davies, K.M, 1998. Identification of flavonol and anthocyanin metabolites in leaves of Petunia "Mitchell" and its Lc transgenic. Phytochemistry 49: 1427–1430.
Bovy, A. R. de Vos, Kemper, M, Schijien, E., Almenar Pertejo, M., Muir, S., Collins, G., Robinson, S., Verhoeyen, M., Hughes, S., Santos-Buelga, C., van Tunen, A.,2002.High-Flavonol Tomatoes Resulting from the Heterologous Expression of the Maize Transcription Factor Genes LC and C1. The Plant Cell, 14: 2509-2526.
Chang, S., Puryear, J., Cairney, J., 1993.A simple and efficient method for isolating RNA from pine trees. Plant Mol.Biol.Rep., 11:113-116.
Chu, C.C., Proc.Sym.Plant tissue culture 1978. Science Press, Peking: pages 45-50.
Dellaporta, S.L., Greenblatt, I.M., Kermicle, J.L., 1988.Molecular cloning of the R-nj gene by transposon tagging with Ac. In Chromosome Structure and Function: impact of new concept, J.P.Gustafson and R.Appels, eds(New York: Plenum Press), pp.263-282.
Goodrich,J.,Carpenter, R.,1992.A common gene regulates pigmentation pattern in diverse plant species. Cell 68, 955-964.
Gruber, M. y., H. Ray, 2002. Novel regulatory genes involved in condensed tannins synthesis in plants. US. Patent No# 2000022156.
Hagerman, A.E., Butler, L.G., 1994. Assay of condensed tannins or flavonoid oligomers and related flavonoids in plants. Meth Enz , 234: 429-437.
Hellens, R., Mullineaux, P., Klee, H. 2000 A guide to Agrobacterium binary Ti vectors.Trends in Plant Sci. 5 : 446-451.
Hesamzadeh Hejazi ,S.M., Abde-Mishani, S., Arcioni , S., Alizadeh, H.,  Tavakol Afshar, R. , Hosseinzadeh, A., 2004.Production of bloat safe alfalfa plants through genetic transformation.Ph.D thesis, University of Tehran, IR.
Holton, T.A., Edwina, C.C., 1995.Genetics and biochemistry of anthocyanin biosynthesis.The Plant Cell, 7: 1071-1083.
Jende-Strid, J. 1993. Genetic control of flavonoid biosynthesis in barley. Hereditas. 119: 187-204.
Johnson, L.B., Schlarbaum, S.,E., Skinner, D.Z., 1984.Variation in phenotype and chromosome number in alfalfa protoclones regenerated from nonmutagenized  calli, Crop. Sci., 24: 948-952.
Joseph, R., Tanner, G. and Larkbl, P.,1998. Proantbocyanidin synthesis in the forage legume Onobrychis viciifolia -a study of chalcone synthase. Dihydroflavonol 4-reductase and leucoantho cyanidin 4- reductase in developing leaves. Aust. J. Plant Physiol. 25: 271-278.
Kamate, K., Rodriguez- Llorente, I.D., Schotle, M., Durand, P., Ratet, P., Kondorosi, E., Kondorosi, A., Trinh, T.H.,2000. Transformation of floral organs with GFP in Medicago Truncatula .Plant Cell Rep.19: 647-653.
Koornneef, M., Dellaert, L.W.M. and van der Veen, J.H. 1982a.EMS and radiation-induced mutation frequencies at individual loci in Arabidopsis thaliana (L.) Heynh. Mutation Res. 93:109-123.
Koornneef, M., Luiten, W., de Vlaming, P. and Schram, A.W. 1982b. A gene controlling flavonoid-3'-hydroxylation in Arabidopsis. Arab. Inf. Serv. 19:113-115.
Li, Y.G., Tanner, G.J., Delves, A.C., Larkin, P.J., 1993.Symetric somatic hybrid plants between Medicago sativa L. and Onobrychis vicifolia Scop.Theor.Appl.Genet. 87: 455-463.
Li, Y.G. Tanner, G.J., Larkin, P.J., 1996. The DMACA-HCL protocol and the threshold proantho cyanidin content for bloat safety in forage legumes. J. Sci. Food Agric. 70: 89-101.
Ludwig, S.R., Habera, L.F., 1989. Lc, a member of the maize R gene family responsible for tissue-specific anthocyanin production, encodes a protein similar to transcriptional activators and contains the myc-homology region .Proc.Natl.Acad.Sci. USA 86, 7092-7096.
Marescalchi, O., Seali, V., 1998. Flow-Cytometric analysis of intraspecific genome size variations in Bacillus atticus (insecta, Phasmatodea). Genome, 41 : 629-635.
Quattrocchio, F., Wing, J.F., Leppen, H.T.C., Mol, J.N.M.,1993.regulatory genes controlling anthocyanin pigmentation are functionally conserved among plant species and have distinct sets of target genes. Plant Cell 5, 1497-1512.
Rob,J. A., Barry, T.N. , McNabb, W.c.,2000. Review paper for polyphenols and agriculture :beneficial effects of proanthocyanidins in forage.Agri.Ecosystems and Environment, 75 : 1-12.
Rumbaugh, M.D., 1985. Breeding bloat-safe cultivars of bloat-causing legumes, in:Barnes, R.F., Ball, P.R., Bringham, R.w.,(Eds.) Forage legumes for Energy-Efficient Animal  Production.USDA, Washington.Proc.Bilaternal Workshop, Plamerston North, NZ, April 1984, pp.238-245.
Sambrook, J., Fritsch, E f., Maniatis, T., 1989. Molecular cloning : A laboratory manual-  Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, ed. 2nd.
Scalbert, A.1991.Antimicrobial properties of tannins.Phytochemistry 30: 3875-3883.
Schenk, R. U. and Hildebrant, A. C., 1972. Medium and techniques for induction and growth of monocotyledonous and dicotyledonous plant cell cultures.Can. J. Bot. 50:199-204.
Susan Marles, M.A., Ray, H., Gruber, M. Y., 2003.New perspectives on proanthocyanidin biochemistry and molecular regulation Phytochemistry 64 :367-383.
Sylvie, A., Delseny, M., Devic, M., 1997. BANYULS, a novel negative regulatorof flavonoid biosynthesis in the Arabidopsis seed coat. The Plant J.11(2): 289-299.
Szymanski, D.B., Klis, D.A., Larkin, J.C., Marks, M.D., 1998. A regulator of  Arabidopsis trichome initiation. Genetics. 149: 565-77.
Tanner, G.J., Joseph, R.G., Li, Y.G., Larkin, P.J., 1997.Towards bloat safe pastures, Feedmix, University of Cambridge. UK.
Tonelli, C., Consonni, G., Dolfini, S.F.,1991. Genetic and molecular analysis of Sn, a light–inducible, tissue specific regulatory gene in maize. Mol.Gen.Genet. 225: 401-410.
Trieu, A.,T., S.H., Burleigh, I., V., Kardailsky, M.J., Hrrison, 2000.Technical advance :transformation of Medicago sp. via infiltration of seedling of flowering plants with agrobacterium. Plant J., 22: 531-541.
Walker Peach, CR, Velten, J., 1994.Agrobactrium-mediated gene transfer to plant cell:cointegrate and binary vector systems in :Gelvin Sb, Schilperoot RA(eds) Plants Molecular Biology Manual.Kluwer, Dordrecht, pp.B1-B19.
Winkel-Shirley, B., 2001.Flavonoid biosynthesis.A colorful model for genetic ,biochemistry,cell biology, and biotechnology. Plant Physiol., 126: 485-493.
Iranian Journal of Rangelands and Forests Plant Breeding and Genetic Research
Volume 13, Issue 3 - Serial Number 21
October 2006
Pages 295-338

Files

Share

How to cite

Statistics