Functional Characterization of Genes Involved in Legume Nodulation Using Hairy Root Cultures

  • Jawahar Singh
  • Kamal Kumar
  • Praveen Kumar VermaEmail author
Part of the Rhizosphere Biology book series (RHBIO)


Legumes, the second most important crop to humans possess unique ability to fix atmospheric nitrogen, making them one of the major contributors to sustainable agriculture. In legumes, molecular characterization of genes by stable transformation is difficult due to their recalcitrant nature to the whole-plant regeneration in desired varieties. The Agrobacterium rhizogenes-mediated generation of transgenic hairy roots or composite plants may facilitate a rapid and convenient alternative to study nodule biology. Functional analysis of genes involved in legume nodulation has been proven as successful for model legumes, viz., Medicago truncatula and Lotus japonicus, using transgenic hairy roots. Besides sharing some common features of nodulation among legumes, the symbiotic signaling is a complex and specific process. Here, we describe an improved protocol for hairy root transformation of a legume crop chickpea (Cicer arietinum L.) and the method to study nodulation to uncover the signaling components. Using the described protocol, transgenic hairy roots were generated in chickpea and selected based on the red fluorescence protein (RFP) microscopy. This protocol can be extended to other underutilized legumes.


Chickpea Hairy root Agrobacterium rhizogenes Mesorhizobium ciceri Nodulation 



The authors thank Dr. Jean-Michel Ané, University of Wisconsin, Madison, USA for providing the Agrobacterium rhizogenes ARqua1strain; Dr. Saikat Bhattacharjee, Regional Centre for Biotechnology (RCB), India for providing pSITE-4NB destination vector; and Dr. Solange Oliveira, Universidade de Evora, Evora, Portugal for providing the Mesorhizobium ciceri LMS-1. This work is partially supported by core grant of National Institute of Plant Genome Research and DBT-CPCFG project grant (BT/AGR/CG-Phase II/01/2014). JS acknowledge Council of Scientific and Industrial Research, India for a fellowship.


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Copyright information

© Springer Nature Singapore Pte Ltd. 2020

Authors and Affiliations

  • Jawahar Singh
    • 1
  • Kamal Kumar
    • 1
  • Praveen Kumar Verma
    • 1
    Email author
  1. 1.Plant Immunity LaboratoryNational Institute of Plant Genome ResearchNew DelhiIndia

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