Plant Cell Reports

, Volume 27, Issue 3, pp 471–478 | Cite as

High-efficiency Agrobacterium-mediated transformation of Brachypodium distachyon inbred line Bd21-3

  • John VogelEmail author
  • Theresa Hill
Genetic Transformation and Hybridization


Brachypodium distachyon (Brachypodium) is a small grass with biological attributes (rapid generation time, small genome, diploid accessions, small stature and simple growth requirements) that make it suitable for use as a model system. In addition, a growing list of genomic resources have been developed or are currently under development including: cDNA libraries, BAC libraries, EST sequences, BAC end sequences, a physical map, genetic markers, a linkage map and, most importantly, the complete genome sequence. To maximize the utility of Brachypodium as a model grass it is necessary to develop an efficient Agrobacterium-mediated transformation system. In this report we describe the identification of a transformable inbred diploid line, Bd21-3, and the development of a transformation method with transformation efficiencies as high as 41% of co-cultivated calluses producing transgenic plants. Conducting the co-cultivation step under desiccating conditions produced the greatest improvement in transformation efficiency.


Brachypodium Transformation Agrobacterium Biofuel Model system 



Callus inducing media


Embryogenic callus


Expressed sequence tag


Bacterial artificial chromosome


2,4-Dichlorophenoxyacetic acid


Linsmaier and Skoog basal medium


Murashige and Skoog salts and vitamins



We would like to thank David Garvin for seeds of PI 254867, Roger Thilmony for vector pGPro2, James Thomson for vectors p#1, p#4, p#6 and Naxin Huo for technical assistance. This work was supported by USDA CRIS project 5325-21000-013-00 “Biotechnological Enhancement of Energy Crops.”


  1. Arumuganathan K, Earle E (1991) Estimation of nuclear DNA content of plants by flow cytometry. Plant Mol Biol Rep 9:229–241Google Scholar
  2. Ausubel F, Brent R, Kingston R, Moore D, Seidman J, Smith J, Struhl K (1996) Current protocols in molecular biology. Wiley, New YorkGoogle Scholar
  3. Bablak P, Draper J, Davey M, Lynch P (1995) Plant regeneration and micropropagation of Brachypodium distachyon. Plant Cell Tissue Organ Cult 42:97–107CrossRefGoogle Scholar
  4. Bennett M, Leitch I (2005) Nuclear DNA amounts in Angiosperms: progress, problems and prospects. Ann Bot 95:45–90PubMedCrossRefGoogle Scholar
  5. Cheng M, Hu T, Layton J, Liu C, Fry J (2003) Desiccation of plant tissues post-Agrobacterium infection enhances T-DNA delivery and increases stable transformation efficiency in wheat. In Vitro Cell Dev Biol Plant 39:595–604CrossRefGoogle Scholar
  6. Christiansen P, Didion T, Andersen C, Folling M, Nielsen K (2005) A rapid and efficient transformation protocol for the grass Brachypodium distachyon. Plant Cell Rep 23:751–758PubMedCrossRefGoogle Scholar
  7. DOE (ed) (2006) Breaking the biological barriers to cellulosic ethanol: a Joint Research Agenda. U.S. Department of Energy, Office of Science and Office of Energy EfficiencyGoogle Scholar
  8. Draper J, Mur L, Jenkins G, Ghosh-Biswas G, Bablak P, Hasterok R, Routledge A (2001) Brachypodium distachyon. A new model system for functional genomics in grasses. Plant Physiol 127:1539–1555PubMedCrossRefGoogle Scholar
  9. Feldmann K (1991) T-DNA insertion mutagenesis in Arabidopsis: mutational spectrum. Plant J 1:71–82CrossRefGoogle Scholar
  10. Hasterok R, Marasek A, Donnison I, Armstead I, Thomas A, King I, Wolny E, Idziak D, Draper J, Jenkins G (2006) Alignment of the genomes of Brachypodium distachyon and temperate cereals and grasses using bacterial artificial chromosome landing with fluorescence in situ hybridization. Genetics 173:349–362PubMedCrossRefGoogle Scholar
  11. Huo N, Gu Y, Lazo G, Vogel J, Coleman-Derr D, Luo M, Thilmony R, Garvin D, Anderson O (2006) Construction and characterization of two BAC libraries from Brachypodium distachyon, a new model for grass genomics. Genome 49:1099–1108PubMedCrossRefGoogle Scholar
  12. Huo N, Lazo G, Vogel J, You F, Ma Y, Hayden D, Colemann-Derr D, Hill T, Dvorak J, Anderson O, Luo M, Gu Y (2007) The nuclear genome of Brachypodium distachyon: analysis of BAC end sequences. Funct Integr Genomics (in press)Google Scholar
  13. Jeon J, Lee S, Jung K, Jun S, Jeong D, Lee J, Kim C, Jang S, Lee S, Yang K, Nam J, An K, Han M, Sung R, Choi H, Yu J, Choi J, Cho S, Cha S, Kim S, An G (2000) T-DNA insertional mutagenesis for functional genomics in rice. Plant J 22:561–570PubMedCrossRefGoogle Scholar
  14. Kohli A, Twyman R, Abranches R, Wegel E, Stoger E, Christou P (2003) Transgene integration, organization and interaction in plants. Plant Mol Biol 52:247–258PubMedCrossRefGoogle Scholar
  15. Lazo G, Stein P, Ludwig R (1991) A DNA transformation-competent Arabidopsis genomic library in Agrobacterium. Biotechnology 9:963–967PubMedCrossRefGoogle Scholar
  16. Svitashev S, Somers D (2002) Characterization of transgene loci in plants using FISH: a picture is worth a thousand words. Plant Cell Tissue Organ Cult 69:205–214CrossRefGoogle Scholar
  17. Thilmony R, Guttman M, Chiniquy D, Blechl A (2006) pGPro1, a novel binary vector for monocot promoter characterization. Plant Mol Biol Rep 24:57–69CrossRefGoogle Scholar
  18. Tyagi A, Mohanty A (2000) Rice transformation for crop improvement and functional genomics. Plant Sci 158:1–18PubMedCrossRefGoogle Scholar
  19. Vogel J, Gu Y, Twigg P, Lazo G, Laudencia-Chingcuanco D, Hayden D, Donze T, Vivian L., Stamova B, Coleman-Derr D (2006a) EST sequencing and phylogenetic analysis of the model grass Brachypodium distachyon. Theor Appl Genet 113:186–195PubMedCrossRefGoogle Scholar
  20. Vogel J, Garvin D, Leong O, Hayden D (2006b) Agrobacterium-mediated transformation and inbred line development in the model grass Brachypodium distachyon. Plant Cell Tiss Org Cult 85:199–211Google Scholar
  21. Wang M, Li Z, Matthews P, Upadhyaya N, Waterhouse P (1998) Improved vector for Agrobacterium tumefaciens-mediated transformation of monocot plants. Acta Hortic 461:401–408Google Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  1. 1.USDA Western Regional Research CenterAlbanyUSA

Personalised recommendations