Advertisement

Maize Transformation Using the Morphogenic Genes Baby Boom and Wuschel2

  • Todd Jones
  • Keith Lowe
  • George Hoerster
  • Ajith Anand
  • Emily Wu
  • Ning Wang
  • Maren Arling
  • Brian Lenderts
  • William Gordon-Kamm
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1864)

Abstract

Despite the fact that maize transformation has been available for over 25 years, the technology has remained too specialized, labor-intensive, and inefficient to be useful for the majority of academic labs. Compounding this problem, future demands in maize genome engineering will likely require a step change beyond what researchers view as “traditional” maize transformation methods. Recently, we published on our use of constitutively expressed morphogenic transcription factors Baby Boom (Bbm) and Wuschel2 (Wus2) to improve maize transformation, which requires CRE-mediated excision before regeneration of healthy, fertile T0 plants. Moving beyond this first-generation system, we have developed a new expression system for Bbm and Wus2, using a non-constitutive maize phospholipid transferase protein promoter (Pltppro) driving Bbm expression and a maize auxin-inducible promoter (Axig1pro) for WUS2 expression. Using this combination of expression cassettes, abundant somatic embryos rapidly form on the scutella of Agrobacterium-transformed zygotic immature embryos. These somatic embryos are uniformly transformed and can be directly germinated into plants without a callus phase. Transformed plants are sent to the greenhouse in as little as 1 month, and these T0 plants match the seed-derived phenotype for the inbred and are fertile. T1 seeds germinate normally and have a uniformly wild-type inbred phenotype. This new system represents a rapid, user-friendly transformation process that can potentially facilitate high-throughput production of transgenic T0 plants in B73, Mo17, and the recently developed Fast-Flowering Mini-Maize.

Key words

Maize Transformation Bbm Wus2 PLTPpro Axig1pro Morphogenic genes 

References

  1. 1.
    Songstad DD, Armstrong CL, Peterson WL, Hairston B, Hinchee MAW (1996) Production of transgenic maize plants and progeny by bombardment of Hi-II immature embryos. In Vitro Cell Dev Biol Plant 32:179–183CrossRefGoogle Scholar
  2. 2.
    Lowe K, Wu E, Wang N, Hoerster G, Hastings C, Cho M-J, Scelonge C, Lenderts B, Chamberlin M, Cushatt J, Wang L, Ryan L, Khan T, Chow-Yiu J, Hua W, Yu M, Bahn J, Bao Z, Brink K, Igo E, Rudrappa B, Shamseer PM, Bruce W, Newman L, Shen B, Zheng P, Bidney D, Falco C, Register J, Zhao Z-Y, Xu D, Jones T, Gordon-Kamm W (2016) Morphogenic regulators Baby boom and Wuschel improve monocot transformation. Plant Cell 28:1–19CrossRefGoogle Scholar
  3. 3.
    Lowe K, La Rota M, Hoerster G, Hastings C, Wang N, Chamberlin M, Wu E, Jones T, Gordon-Kamm W (2018) Rapid genotype independent maize transformation via direct somatic embryogenesis. In Vitro Cell Dev Biol Plant 54(3):240–252CrossRefGoogle Scholar
  4. 4.
    Garnaat C, Lowe K, Roth B (2002) Zm-AXIG1-specific polynucleotides and methods of use. WO2002006499Google Scholar
  5. 5.
    McCaw ME, Wallace JG, Albert PS, Buckler ES, Birchler JA (2016) Fast-flowering mini-maize: seed to seed in 60 days. Genetics 204:35.  https://doi.org/10.1534/genetics.116.191726 CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Anand A, Bass SH, Cho H-J, Klein TM, Lassner M, McBride KE (2017) Methods and compositions of improved plant transformation. PCT/US2016/049132Google Scholar
  7. 7.
    Yang H, Schmuke JJ, Flagg LM, Roberts JK, Allen EM, Ivashuta S, Gilbertson LA, Armstrong TA, Christian AT (2009) A novel real-time polymerase chain reaction method for high throughput quantification of small regulatory RNAs. Plant Biotechnol J 7:621–630CrossRefGoogle Scholar
  8. 8.
    Truett GE, Heeger P, Mynatt RL, Truett AA, Walker JA, Warmman ML (2000) Preparation of PCR-quality mouse genomic DNA with hot sodium hydroxide and tris (HotSHOT). BioTechniques 29:52–54CrossRefGoogle Scholar
  9. 9.
    Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods 25:402–408CrossRefGoogle Scholar
  10. 10.
    Boutilier K, Offringa R, Sharma VK, Kieft H, Ouellet T, Zhang L, Hattori J, Liu CM, van Lammeren AA, Miki BL, Custers JB, van Lookeren Campagne MM (2002) Ectopic expression of BABY BOOM triggers a conversion from vegetative to embryonic growth. Plant Cell 14:1737–1749CrossRefGoogle Scholar
  11. 11.
    Gordon-Kamm WJ, Helentjaris TG, Lowe KS, Shen B, Tarczynski MC Zheng P (2005) Ap2 domain transcription factor Odp2 (ovule development protein 2) and methods of use. Patent no. WO2005/075655-A2Google Scholar
  12. 12.
    Laux T, Mayer KF, Berger J, Jürgens G (1996) The WUSCHEL gene is required for shoot and floral meristem integrity in Arabidopsis. Development 122:87–96PubMedGoogle Scholar
  13. 13.
    Gallois JL, Woodward C, Reddy GV, Sablowski R (2002) Combined SHOOT MERISTEMLESS and WUSCHEL trigger ectopic organogenesis in Arabidopsis. Development 129:3207–3217PubMedGoogle Scholar
  14. 14.
    Lowe KS, Cahoon RE, Scelong CJ, Tao Y, Gordon-Kamm WJ, Bruce WB, Newman LJ (2007) Wuschel (WUS) Gene Homologs Patent No US7256322Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Todd Jones
    • 1
  • Keith Lowe
    • 1
  • George Hoerster
    • 1
  • Ajith Anand
    • 1
  • Emily Wu
    • 1
  • Ning Wang
    • 1
  • Maren Arling
    • 1
  • Brian Lenderts
    • 1
  • William Gordon-Kamm
    • 1
  1. 1.Corteva Agriscience™Agriculture Division of DowDuPont™JohnstonUSA

Personalised recommendations