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Methods for Estimation of Nitrogen Components in Plants and Microorganisms

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Nitrogen Metabolism in Plants

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2057))

Abstract

Nitrogen (N2) is the most necessary element in the atmosphere, it is an energetic micronutrient for plant growth and development after water, besides its key role in chlorophyll production, which is crucial for photosynthesis process. Biological nitrogen fixation is measured to be the most potent method to deliver a fixed way of nitrogen to the plants. Plant depends on free-living and symbiotic microbes present in the soil for nitrogen because it cannot be absorbed by the plant itself directly from the atmosphere. Many techniques were reported in the laboratory for nitrogen estimation till now, but Kjeldahl digestion and acetylene reduction assay (ARA) techniques became the most popular. In this chapter, we focus on the most common and popular methods used to determine plant N2; awareness obtained through the wide application of these methods should offer the source for the N2 fixation rate in agriculture system.

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References

  1. Shin W, Islam R, Benson A, Joe MM, Kim K, Gopal S, Samaddar S, Banerjee S, Sa T (2016) Role of Diazotrophic Bacteria in biological nitrogen fixation and plant growth improvement. Korean J Soil Sci Fert 49:17–29

    Article  CAS  Google Scholar 

  2. Figueiredo MVB, Mergulhao ACES, Sobral JK, Junior MAL, Araujo A (2013) Biological nitrogen fixation: importance, associated diversity, and estimates. In: Arora NK (ed.) Plant microbe symbiosis: fundamentals and advances. https://doi.org/10.1007/978-81-322-1287-4_10, © Springer India

  3. Franche C, Lindstro MK, Elmerich C (2009) Nitrogen-fixing bacteria associated with leguminous and non-leguminous plants. Plant Soil 321:35–59

    Article  CAS  Google Scholar 

  4. Canfield D, Glazer AN, Falkowski PD (2010) The evolution and future of earth’s nitrogen cycle. Science 330:192–196

    Article  CAS  Google Scholar 

  5. Zehr JP, Jenkins BD, Short SM, Steward GF (2003) Nitrogenase gene diversity and microbial community structure: a cross-system comparison. Environ Microbiol 5:539–554

    Article  CAS  Google Scholar 

  6. Falkowski PG, Fenchel T, Delong EF (2008) The microbial engines that drive Earth’s biogeochemical cycles. Science 320:1034–1039

    Article  CAS  Google Scholar 

  7. Hardy RWF, Knight E Jr (1968) The biochemistry and postulated mechanisms of N2 fixation. In: Reinhold L (ed) Progress in phytochemistry. John Wiley and Sons Ltd, Sussex, England, pp 387–469

    Google Scholar 

  8. Burris RH, Wilson PW (1957) Methods for measurement of nitrogen fixation. In: Colowick SP, Kaplan NO (eds) Methods in enzymology, vol 4. Academic Press, New York, pp 355–367

    Google Scholar 

  9. Campbell NE, Dular R, Lees H (1967) The production of 13N2 by 50 Mev protons for use in biological nitrogen fixation. Can J Microbiol 13:587–599

    Article  CAS  Google Scholar 

  10. Nicholas DJD, Silvester DJ, Fowle JF (1961) Use of radioactive nitrogen in studying nitrogen fixation in bacterial cells and their extracts. Nature 189:634–636

    Article  CAS  Google Scholar 

  11. Carnahan JE, Mortenson LE, Mower HF, Castle JE (1960) Nitrogen fixation in cell-free extracts of Clostridium pasteurianum. Biochim Biophys Acta 44:520–535

    Article  CAS  Google Scholar 

  12. Mortenson LE (1961) A simple method for measuring nitrogen fixation by cell-free enzyme preparations of Clostridium pasteurianum. Anal Biochem 2:216–220

    Article  CAS  Google Scholar 

  13. Dilworth MJ, Subramanian D, Munson TO, Burris RH (1965) The adenosine triphosphate requirement for nitrogen fixation in cell-free extracts of Clostridium pastcurianum. Biochim Biophys Acta 99:486–503

    Article  CAS  Google Scholar 

  14. Mortenson LE (1964) Ferredoxin and ATP, requirements for nitrogen fixation in cell-free extracts of Clostridium pasteurianum. Proc Natl Acad Sci U S A 52:272–279

    Article  CAS  Google Scholar 

  15. Kjeldahl J (1883) Neue methode zur bestimmung des stickstoffs in organischen körpern. Fresenius J Anal Chem 22:366–382

    Article  Google Scholar 

  16. Labconco CA (1998) Guide to Kjeldahl nitrogen determination methods and apparatus. Labconco cCorporation, Houston

    Google Scholar 

  17. Dumas JBA (1831) Procedes de l’analyse organic. Ann Chim Phys 247:198–213

    Google Scholar 

  18. Stem WR (1993) Nitrogen fixation and transfer in intercrop systems. Field Crop Res 34:335–356

    Article  Google Scholar 

  19. Dilworth MJ (1966) Acetylene reduction by nitrogen-fixing preparations from Clostridium pasteurianum. Biochem Biophys Acta 127:285–294

    Article  CAS  Google Scholar 

  20. Crews TE, Kurina LM, Vitousek PM (2001) Organic matter and nitrogen accumulation and nitrogen fixation during early ecosystem development in Hawaii. Biogeochemistry 52:259–279

    Article  CAS  Google Scholar 

  21. Castle S (2010) Acetylene reduction assay (ARA): measuring Nitrogenase activity. Aridlands ecology lab protocol modified: 2010.02.16

    Google Scholar 

  22. Das S, De TK (2018) Microbial assay of N2 fixation rate, a simple alternate for acetylene reduction assay. MethodsX 5:909–914

    Article  Google Scholar 

  23. Unkovich M, Herridge D, Peoples M, Cadisch G, Boddey B, Giller K, Alves B, Chalk P (2008) Measuring plant-associated nitrogen fixation in agricultural systems. Australian Centre for International Agricultural Research (ACIAR), Canberra, ACT, Australia, pp 45–62

    Google Scholar 

  24. Lee D, Nguyen V, Littlefield S (1996) Comparison of methods for determination of nitrogen levels in soil, plant and body tissues, and water. Commun Soil Sci Plant Anal 27:83–793

    Google Scholar 

  25. Kayasth M, Gera R, Dudeja SS, Sharma PK, Kumar V (2014) Studies on salinization in Haryana soils on free-living nitrogen-fixing bacterial populations and their activity. J Basic Microbiol 54:170–179

    Article  CAS  Google Scholar 

  26. Lee KK, Watanabe I (1977) Problems of the acetylene reduction technique applied to water- saturated paddy soils. Appl Environ Microbiol 34:654–660

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Portillo MC, Leff JW, Lauber CL, Fierer N (2013) Cell size distributions of soil bacterial and archaeal taxa. Appl Environ Microbiol 79:7610–7617

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Key Laboratory of Sugarcane Biotechnology and Genetic Improvement (Guangxi), Ministry of Agriculture, Sugarcane Research Center, Chinese Academy of Agricultural Sciences, Guangxi key Laboratory of Sugarcane Genetic Improvement, Sugarcane Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, Guang, China

    Pratiksha Singh, Rajesh Kumar Singh, Qi-Qi Song, Hai-Bi Li, Li-Tao Yang & Yang-Rui Li

  2. College of Agriculture, Guangxi University, Nanning, Guang, China

    Pratiksha Singh, Rajesh Kumar Singh, Qi-Qi Song, Hai-Bi Li, Li-Tao Yang & Yang-Rui Li

  3. Guangxi key Laboratory of Crop Genetic Improvement and Biotechnology, Nanning, Guang, China

    Pratiksha Singh & Rajesh Kumar Singh

Authors
  1. Pratiksha Singh
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  2. Rajesh Kumar Singh
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  3. Qi-Qi Song
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  4. Hai-Bi Li
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  5. Li-Tao Yang
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  6. Yang-Rui Li
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Editor information

Editors and Affiliations

  1. National Institute of Plant Genome Research, New Delhi, India

    Kapuganti Jagadis Gupta

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Singh, P., Singh, R.K., Song, QQ., Li, HB., Yang, LT., Li, YR. (2020). Methods for Estimation of Nitrogen Components in Plants and Microorganisms. In: Gupta, K. (eds) Nitrogen Metabolism in Plants. Methods in Molecular Biology, vol 2057. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9790-9_10

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  • DOI: https://doi.org/10.1007/978-1-4939-9790-9_10

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9789-3

  • Online ISBN: 978-1-4939-9790-9

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