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High-Resolution Cell-Type Specific Analysis of Cytokinins in Sorted Root Cell Populations of Arabidopsis thaliana

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Plant Hormones

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1497))

Abstract

We describe a method combining fluorescence-activated cell sorting (FACS) with one-step miniaturized isolation and accurate quantification of cytokinins (CKs) using ultra-high performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS) to measure these phytohormones in specific cell types of Arabidopsis thaliana roots. The methodology provides information of unprecedented resolution about spatial distributions of CKs, and thus should facilitate attempts to elucidate regulatory networks involved in root developmental processes.

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References

  1. Petersson SV, Johansson AI, Kowalczyk M et al (2009) An auxin gradient and maximum in the Arabidopsis root apex shown by high-resolution cell-specific analysis of IAA distribution and synthesis. Plant Cell 21:1659–1668

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Antoniadi I, Plačková L, Simonovik B et al (2015) Cell-type specific cytokinin distribution within the Arabidopsis primary root apex. Plant Cell 27:1955–1967

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Birnbaum K, Shasha DE, Wang JY et al (2003) A gene expression map of the Arabidopsis root. Science 302:1956–1960

    Article  CAS  PubMed  Google Scholar 

  4. Yadav RK, Girke T, Pasala S et al (2009) Gene expression map of the Arabidopsis shoot apical meristem stem cell niche. Proc Natl Acad Sci U S A 106:4941–4946

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Pěnčík A, Simonovik B, Petersson SV et al (2013) Regulation of auxin homeostasis and gradients in Arabidopsis roots through the formation of the IAA catabolite oxIAA. Plant Cell 25:3858–3870

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Petersson SV, Lindén P, Moritz T et al (2015) Cell-type specific metabolic profiling of Arabidopsis thaliana protoplasts as a tool for plant systems biology. Metabolomics 11:1679–1689

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Tarkowská D, Novák O, Floková K et al (2014) Quo vadis plant hormone analysis? Planta 240:55–76

    Article  CAS  PubMed  Google Scholar 

  8. Svačinová J, Novák O, Plačková L et al (2012) A new approach for cytokinin isolation from Arabidopsis tissues using miniaturized purification: pipette tip solid-phase extraction. Plant Methods 8:17

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Novák O, Hauserová E, Amakorová P et al (2008) Cytokinin profiling in plant tissues using ultra-performance liquid chromatography–electrospray tandem mass spectrometry. Phytochemistry 69:2214–2224

    Article  CAS  PubMed  Google Scholar 

  10. Liu Z, Cai BD, Feng YQ (2012) Rapid determination of endogenous cytokinins in plant samples by combination of magnetic solid phase extraction with hydrophilic interaction chromatography-tandem mass spectrometry. J Chromatogr B 891–892:27–25

    Article  CAS  Google Scholar 

  11. Du FY, Ruan GH, Liang SH et al (2012) Monolithic molecularly imprinted solid-phase extraction for the selective determination of trace cytokinins in plant samples with liquid chromatography-electrospray tandem mass spectrometry. Anal Bioanal Chem 404:489–501

    Article  CAS  PubMed  Google Scholar 

  12. Rappsilber J, Mann M, Ishihama Y (2007) Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips. Nat Protoc 2:1896–1906

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by the Ministry of Education, Youth and Sports of the Czech Republic (the National Program for Sustainability I No. LO1204 and the “Návrat” program LK21306) and by the Czech Science Foundation (No. GA14-34792S), the European Molecular Biology Organization (EMBO ASTF 297-2013), the Erasmus program (European Commission: Lifelong Learning Programme), Development-The Company of Biologists (DEVTF2012), the Plant Fellows programme (http://www.plantfellows.ch/), the Swedish Governmental Agency for Innovation Systems (VINNOVA), and the Swedish Research Council (VR).

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Correspondence to Ondřej Novák .

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Novák, O., Antoniadi, I., Ljung, K. (2017). High-Resolution Cell-Type Specific Analysis of Cytokinins in Sorted Root Cell Populations of Arabidopsis thaliana . In: Kleine-Vehn, J., Sauer, M. (eds) Plant Hormones. Methods in Molecular Biology, vol 1497. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6469-7_19

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  • DOI: https://doi.org/10.1007/978-1-4939-6469-7_19

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6467-3

  • Online ISBN: 978-1-4939-6469-7

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