Functional Characterization of Genes Involved in Legume Nodulation Using Hairy Root Cultures
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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.
KeywordsChickpea 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.
- Atlas RM (1997) In: Parks LC (ed) Handbook of microbial media, 2nd edn. CRC Press Inc, Boca Raton, 401 pGoogle Scholar
- Chakrabarty R, Banerjee R, Chung MS, Farman M, Citovsky V, Hogenhout SA, Tzfira T, Goodin M (2007) pSITE vectors for stable integration or transient expression of autofluorescent protein fusions in plants: probing Nicotiana benthamiana–virus interactions. Mol Plant Microbe Interact 20:740–750CrossRefPubMedGoogle Scholar