Abstract
The temperate cereal barley is grown as a source of food, feed, and malt. The development of a broad range of genetic resources and associated technologies in this species has helped to establish barley as the prime model for the other Triticeae cereals. The specific advantage of the transformation method presented here is that transgene homozygosity is attained in the same generation as the transgenic event occurred through the coupling of haploid technology with Agrobacterium-mediated transformation. Pollen is haploid and, following transformation, can be induced to regenerate into haploid plantlets, which can subsequently subjected to colchicine treatment to obtain diploid, genetically fixed plants. The routine application of the method based on the winter-type barley cultivar ‘Igri’ over a period of over 10 years has achieved an average yield of about two transgenic plants per donor spike. The whole procedure from pollen isolation to non-segregating transgenic, mature grain takes less than 12 months.
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References
Maluszynski M, Kasha KJ, Szarejko I (2003) Published doubled haploid protocols in plant species. In: Maluszynski M et al (eds) Doubled haploid production in crop plants: a manual. Kluwer Academic Publishers, Dordrecht, The Netherlands, pp 309–335
Bolik M, Koop HU (1991) Identification of embryogenic microspores of barley (Hordeum vulgare L.) by individual selection and culture and their potential for transformation by microinjection. Protoplasma 162:61–68
Olsen L (1991) Isolation and cultivation of embryonic microspores from barley (Hordeum vulgare L.). Hereditas 115:255–266
Kuhlmann U, Foroughi-Wehr B, Graner A, Wenzel G (1991) Improved culture system for microspores of barley to become a target for DNA uptake. Plant Breed 107:165–168
Jähne A, Becker D, Brettschneider R, Lörz H (1994) Regeneration of transgenic, microspore-derived, fertile barley. Theor Appl Genet 89:525–533
Yao QA, Simion E, William M, Krochko J, Kasha KJ (1997) Biolistic transformation of haploid isolated microspores of barley (Hordeum vulgare L.). Genome 40:570–581
Carlson AR, Letarte J, Chen J, Kasha KJ (2001) Visual screening of microspore-derived transgenic barley (Hordeum vulgare L.) with green-fluorescent protein. Plant Cell Rep 20:331–337
Wan Y, Lemaux PG (1994) Generation of large numbers of independently transformed fertile barley plants. Plant Physiol 104:37–48
Kumlehn J, Serazetdinova L, Hensel G, Becker D, Lörz H (2006) Genetic transformation of barley (Hordeum vulgare L.) via infection of androgenetic pollen cultures with Agrobacterium tumefaciens. Plant Biotechnol J 4:251–261
Stein N, Perovic D, Kumlehn J, Pellio B, Stracke S, Streng S, Ordon F, Graner A (2005) The eukaryotic translation initiation factor 4E confers multiallelic recessive Bymovirus resistance in Hordeum vulgare (L.). Plant J 42:912–922
Radchuk V, Borisjuk L, Radchuk R, Steinbiss HH, Rolletschek H, Broeders S, Wobus U (2006) Jekyll encodes a novel protein involved in the sexual reproduction of barley. Plant Cell 18:1652–1666
Gurushidze M, Hensel G, Hiekel S, Schedel S, Valkov V, Kumlehn J (2014) True-breeding targeted gene knock-out in barley using designer TALE-nuclease in haploid cells. PLoS One 9(3):e92046
Baumer M, Cais R (2000) Abstammung der Gerstensorten, Bayerische Landesanstalt für Bodenkultur und Pflanzenbau. 134 pp
Ooms G, Hooykaas PJJ, Van Veen RJM, Van Beelan P, Regensburg-Tuink TJG, Schilperoort RA (1982) Octopine Ti-plasmid deletion mutants of Agrobacterium tumefaciens with emphasis on the right side of the T-region. Plasmid 7:15–29
Hoekema A, Hirsch PR, Hooykaas PJJ, Schilperoort RA (1983) A binary plant vector strategy based on separation of vir and T region of the Agrobacterium tumefaciens Ti-plasmid. Nature 303:179–180
Komari T, Hiei Y, Saito Y, Murai N, Kumashiro T (1996) Vectors carrying two separate T-DNAs for co-transformation of higher plants mediated by Agrobacterium tumefaciens and segregation of transformants free from selection markers. Plant J 10:165–174
Lazo GR, Stein PA, Ludwig RA (1991) A DNA transformation-competent Arabidopsis genomic library in Agrobacterium. Biotechnology 9:963–967
Himmelbach A, Zierold U, Hensel G, Riechen J, Douchkov D, Schweizer P, Kumlehn J (2007) A set of modular binary vectors for the transformation of cereals. Plant Physiol 145: 1192–1200
Kumlehn J (2008) The IPKb vector set: modular binary plasmids for cereal transformation. Information Systems for Biotechnology News Report, Virginia Tech, Blacksburg, April issue, pp 3–6
Acknowledgment
We are grateful to the Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, for providing our research group with excellent working conditions.
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Otto, I., Müller, A., Kumlehn, J. (2015). Barley (Hordeum vulgare L.) Transformation Using Embryogenic Pollen Cultures. In: Wang, K. (eds) Agrobacterium Protocols. Methods in Molecular Biology, vol 1223. Springer, New York, NY. https://doi.org/10.1007/978-1-4939-1695-5_7
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DOI: https://doi.org/10.1007/978-1-4939-1695-5_7
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