Biology and Fertility of Soils

, Volume 52, Issue 2, pp 277–282 | Cite as

Fluorescence resonance energy transfer (FRET)-based technique for tracking of endophytic bacteria in rice roots

  • Avishek Banik
  • Subhra Kanti Mukhopadhaya
  • Animesh Sahana
  • Debasis Das
  • Tushar Kanti DangarEmail author
Short Communication


Visualization of endophytes inside the host tissue has remained a prime interest to the plant microbiologist throughout the last decade. Assessment of a root colonization pattern of a nitrogen-fixing polyvalent plant-growth-promoting endophyte, Azotobacter chroococcum Avi2 (16S rRNA gene NCBI acc. no. KP099933), of Oryza sativa var. Swarna, by a plasmid containing green fluorescent protein (GFP) gene failed because the bacteria formed a cyst on exposure to CaCl2 during the transformation process of the green fluorescent protein gene. A new technique based on fluorescence resonance energy transfer (FRET) was developed to visualize the intracellular location of the bacterium of rice root as an alternative to track the bacterium where the use of GFP is restricted. An Al3+-specific rhodamine-based novel fluorescent ligand (λ ex = 400 nm, λ em = 455 nm), (E)-3′,6′-bis(diethylamino)-2-(2-(pyren-4-ylmethyleneamino)ethyl)spiro[isoindoline-1,9′xanthen]-3-one, was synthesized and used to visualize 100 μM aluminium nitrate treated A. chroococcum inside rice root tissues. Within 6 h of incubation, a clear, stable and distinguishable green fluorescence was observed under the fluorescence microscope form Al3+-treated A. chroococcum colonized in the rice roots when incubated with a ligand. The results confirmed colonization of the bacteria inside the roots.


Rice endophytes Bacterial colonization Fluorescence microscopy FRET 



Financially supported by Indian Council of Agricultural Research (ICAR) project “Application of microorganisms in agriculture and allied sector” (AMAAS) is acknowledged. The University Science Instrumentation Centre (USIC), University of Burdwan, West Bengal, India, for fluorescence microscopy facility is thankfully acknowledged.

Conflict of interest

The authors declare that they have no competing interests.


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Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Avishek Banik
    • 1
  • Subhra Kanti Mukhopadhaya
    • 2
  • Animesh Sahana
    • 3
  • Debasis Das
    • 3
  • Tushar Kanti Dangar
    • 1
    Email author
  1. 1.Microbiology Laboratory, Crop Production Division, ICAR-Central Rice Research InstituteCuttackIndia
  2. 2.Department of MicrobiologyThe University of BurdwanBurdwanIndia
  3. 3.Department of ChemistryThe University of BurdwanBurdwanIndia

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