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An Adapted Protocol for Quantitative Rhizosphere Acidification Assay

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Plant Iron Homeostasis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2665))

Abstract

Acidification of the rhizosphere is a key process in the homeostasis of multiple essential nutrients, including iron. Under iron deficiency, the release of protons from the roots helps solubilize and increase the accessibility of iron in the soil. Rhizosphere acidification has been widely examined in many iron homeostasis studies, generally using a qualitative method based on the color change of bromocresol purple, a pH indicator dye, near the roots. In this chapter, we introduce an adapted version of a rhizosphere acidification assay protocol that allows for the quantitative assessment of small pH changes in the rhizosphere. This colorimetric method also utilizes bromocresol purple, but the ratio of its absorbance at 434 nm and 588 nm is considered to quantify protons released into the assay solution. Furthermore, the assay is compatible with small sample volumes, such as those with young Arabidopsis seedlings.

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References

  1. Houmani H, Rabhi M, Abdelly C, Debez A (2015) Implication of rhizosphere acidification in nutrient uptake by plants: cases of potassium (K), phosphorus (P), and iron (fe). In: Hakeem KR (ed) Crop production and global environmental issues. Springer, Cham, pp 103–122

    Chapter  Google Scholar 

  2. Wang X, Tang C (2018) The role of rhizosphere pH in regulating the rhizosphere priming effect and implications for the availability of soil-derived nitrogen to plants. Ann Bot 121:143–151. https://doi.org/10.1093/aob/mcx138

    Article  CAS  PubMed  Google Scholar 

  3. Santi S, Schmidt W (2009) Dissecting iron deficiency-induced proton extrusion in Arabidopsis roots. New Phytol 183:1072–1084. https://doi.org/10.1111/j.1469-8137.2009.02908.x

    Article  CAS  PubMed  Google Scholar 

  4. Robinson NJ, Procter CM, Connolly EL, Guerinot ML (1999) A ferric-chelate reductase for iron uptake from soils. Nature 397:694–697. https://doi.org/10.1038/17800

    Article  CAS  PubMed  Google Scholar 

  5. Robe K, Izquierdo E, Vignols F et al (2021) The coumarins: secondary metabolites playing a primary role in plant nutrition and health. Trends Plant Sci 26:248–259. https://doi.org/10.1016/j.tplants.2020.10.008

    Article  CAS  PubMed  Google Scholar 

  6. Varotto C, Maiwald D, Pesaresi P et al (2002) The metal ion transporter IRT1 is necessary for iron homeostasis and efficient photosynthesis in Arabidopsis thaliana. Plant J 31:589–599. https://doi.org/10.1046/j.1365-313X.2002.01381.x

    Article  CAS  PubMed  Google Scholar 

  7. Vert G, Grotz N, Dédaldéchamp F et al (2002) IRT1, an Arabidopsis transporter essential for iron uptake from the soil and for plant growth. Plant Cell 14:1223–1233. https://doi.org/10.1105/tpc.001388

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Connolly EL, Fett JP, Guerinot ML (2002) Expression of the IRT1 metal transporter is controlled by metals at the levels of transcript and protein accumulation. Plant Cell 14:1347–1357. https://doi.org/10.1105/tpc.001263

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Grusak MA, Pezeshgi S (1996) Shoot-to-root signal transmission regulates root Fe(III) reductase activity in the dgl mutant of pea. Plant Physiol 110:329–334. https://doi.org/10.1104/pp.110.1.329

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Yi Y, Guerinot ML (1996) Genetic evidence that induction of root Fe(III) chelate reductase activity is necessary for iron uptake under iron deficiency. Plant J 10:835–844. https://doi.org/10.1046/j.1365-313X.1996.10050835.x

    Article  CAS  PubMed  Google Scholar 

  11. Rogers EE, Guerinot ML (2002) FRD3, a member of the multidrug and toxin efflux family, controls iron deficiency responses in Arabidopsis. Plant Cell 14:1787–1799. https://doi.org/10.1105/tpc.001495

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Long TA, Tsukagoshi H, Busch W et al (2010) The bHLH transcription factor POPEYE regulates response to iron deficiency in Arabidopsis roots. Plant Cell 22:2219–2236. https://doi.org/10.1105/tpc.110.074096

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Kim SA, LaCroix IS, Gerber SA, Guerinot ML (2019) The iron deficiency response in Arabidopsis thaliana requires the phosphorylated transcription factor URI. Proc Natl Acad Sci U S A 116:24933–24942. https://doi.org/10.1073/pnas.1916892116

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Yi Y, Saleeba JA, Guerinot ML (1994) Iron uptake in Arabidopsis thaliana. In: Manthey J, Luster D, Crowley DE (eds) Biochemistry of metal micronutrients in the rhizosphere. Lewis Publishers, Boca Raton, pp 295–307

    Google Scholar 

  15. Talone CJ, Gao J, Lynch JR et al (2016) Determination of the ground- and excited-state dipole moments of bromocresol purple in protic and aprotic solvents. Spectrochim Acta A Mol Biomol Spectrosc 156:138–142. https://doi.org/10.1016/j.saa.2015.11.034

    Article  CAS  PubMed  Google Scholar 

  16. Lvov Y, Antipov AA, Mamedov A et al (2001) Urease encapsulation in nanoorganized microshells. Nano Lett 1:125–128. https://doi.org/10.1021/nl0100015

    Article  CAS  Google Scholar 

  17. Schussel LJ, Atwater JE (1995) A urease bioreactor for water reclamation aboard manned spacecraft. Chemosphere 30:985–994. https://doi.org/10.1016/0045-6535(94)00453-2

    Article  CAS  Google Scholar 

  18. Ric de Vos C, Lubberding HJ, Bienfait HF (1986) Rhizosphere acidification as a response to iron deficiency in bean plants on JSTOR. Plant Physiol 81:842–846

    Article  PubMed Central  Google Scholar 

  19. Pizzio GA, Paez-Valencia J, Khadilkar AS et al (2015) Arabidopsis type I proton-pumping pyrophosphatase expresses strongly in phloem, where it is required for pyrophosphate metabolism and photosynthate partitioning. Plant Physiol 167:1541–1553. https://doi.org/10.1104/pp.114.254342

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Rhizosphere Acidification Assay‘ ’ – BIO-PROTOCOL. https://bio-protocol.org/e1676. Accessed 16 May 2019

  21. Pei K, Xiong Y, Li X et al (2018) Colorimetric ELISA with an acid–base indicator for sensitive detection of ochratoxin A in corn samples. Anal Methods 10:30–36. https://doi.org/10.1039/C7AY01959A

    Article  CAS  Google Scholar 

  22. Vengavasi K, Pandey R (2016) Root acidification, a rapid method of screening soybean genotypes for low-phosphorus stress. Ind J Genet Plant Breed 76:213. https://doi.org/10.5958/0975-6906.2016.00025.0

    Article  CAS  Google Scholar 

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Acknowledgments

This work was supported by the Gregory Call Student Research Fund to SO, CM, and KZ, and the National Science Foundation grants (IOS#1754969 and IOS#2143478) to JJ.

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Correspondence to Jeeyon Jeong .

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Omer, S., Macero, C., Daga, D., Zheng, K., Jeong, J. (2023). An Adapted Protocol for Quantitative Rhizosphere Acidification Assay. In: Jeong, J. (eds) Plant Iron Homeostasis. Methods in Molecular Biology, vol 2665. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-3183-6_4

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  • DOI: https://doi.org/10.1007/978-1-0716-3183-6_4

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-3182-9

  • Online ISBN: 978-1-0716-3183-6

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