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    Isolation of cytoskeleton and cytoskeleton-bound polysome, and identification of cytoskeletal proteins from a grain legume
    (Springer Nature Publishing AG, 2026) Kumar, Sunil; Chakraborty, Sohela; Chakraborty, Subhra; Chakraborty, Niranjan
    The plant cytoskeleton is an essential component of cellular architecture, enabling various critical metabolic processes, including cell division, differentiation, expansion and intracellular transport. It consists primarily of three distinct filamentous structures: microtubules, microfilaments and intermediate filaments. These structures are not static; they undergo continuous remodeling in response to environmental signals and developmental cues, which allow plants to adapt to changing conditions. The microtubules and actin filaments have previously been successfully isolated from various plant tissues, contributing to our understanding of their functions. Among the diverse plant families, legumes (Fabaceae) stand out as the third largest, encompassing approximately 20,000 species. They hold significant agricultural importance, ranking second to cereals in global crop production. To fully grasp the developmental and adaptive processes in legumes, it is essential to identify and understand their regulatory components. This chapter focuses on the isolation of cytoskeletal proteins from chickpea, a prominent grain legume, facilitating biochemical and proteomic analyses that may uncover new insights into the functioning of the cytoskeleton in legumes.
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    Quantitative phosphoproteomic analysis of legume using TiO2-based enrichment coupled with isobaric labeling
    (Springer Nature Publishing AG, 2020) Barua, Pragya; Lande, Nilesh Vikram; Kumar, Sunil; Chakraborty, Subhra; Chakraborty, Niranjan
    Phosphorylation of proteins is the most dynamic protein modification, and its analysis aids in determining the functional and regulatory principles of important cellular pathways. The legumes constitute the third largest family of higher plants, Fabaceae, comprising about 20,000 species and are second to cereals in agricultural importance on the basis of global production. Therefore, an understanding of the developmental and adaptive processes of legumes demands identification of their regulatory components. The most crucial signature of the legume family is the symbiotic nitrogen fixation, which makes this fascinating and interesting to investigate phosphorylation events. The research on protein phosphorylation in legumes has been focused primarily on two model species, Medicago truncatula and Lotus japonicus. The development of reciprocal research in other species, particularly the crops, is lagging behind which has limited its beneficial uses in agricultural productivity. In this chapter, we outline the titanium dioxide-based enrichment of phosphopeptides for nuclear proteome analysis of a grain legume, chickpea.
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    Genotype-independent Agrobacterium rhizogenes-mediated root transformation of chickpea: a rapid and efficient method for reverse genetics studies
    (BioMed Central Ltd, 2018) Aggarwal, Pooja Rani; Nag, Papri; Choudhary, Pooja; Chakraborty, Niranjan; Chakraborty, Subhra
    Background: Chickpea (Cicer arietinum L.), an important legume crop is one of the major source of dietary protein. Developing an efcient and reproducible transformation method is imperative to expedite functional genomics studies in this crop. Here, we present an optimized and detailed procedure for Agrobacterium rhizogenes-mediated root transformation of chickpea. Results: Transformation positive roots were obtained on selection medium after two weeks of A. rhizogenes inoculation. Expression of green fuorescent protein further confrmed the success of transformation. We demonstrate that our method adequately transforms chickpea roots at early developmental stage with high efciency. In addition, root transformation was found to be genotype-independent and the efcacy of our protocol was highest in two (Annigiri and JG-62) of the seven tested chickpea genotypes. Next, we present the functional analysis of chickpea hairy roots by expressing Arabidopsis TRANSPARENT TESTA 2 (AtTT2) gene involved in proanthocyanidins biosynthesis. Overexpression of AtTT2 enhanced the level of proanthocyanidins in hairy roots that led to the decreased colonization of fungal pathogen, Fusarium oxysporum. Furthermore, the induction of transgenic roots does not afect functional studies involving infection of roots by fungal pathogen. Conclusions: Transgenic roots expressing genes of interest will be useful in downstream functional characterization using reverse genetics studies. It requires 1 day to perform the root transformation protocol described in this study and the roots expressing transgene can be maintained for 3–4 weeks, providing sufcient time for further functional studies. Overall, the current methodology will greatly facilitate the functional genomics analyses of candidate genes in root-rhizosphere interaction in this recalcitrant but economically important legume crop.
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    Legume proteomics: Progress, prospects and challenges
    (John Wiley & Sons, 2016) Rathi, Divya; Gayen, Dipak; Gayali, Saurabh; Chakraborty, Subhra; Chakraborty, Niranjan
    Legumes are the major sources of food and fodder with strong commercial relevance, and are essential components of agricultural ecosystems owing to their ability to carry out endosymbiotic nitrogen fixation. In recent years, legumes have become one of the major choices of plant research. The legume proteomics is currently represented by more than 100 reference maps and an equal number of stress-responsive proteomes. Among the 48 legumes in the protein databases, most proteomic studies have been accomplished in two model legumes, soybean, and barrel medic. This review highlights recent contributions in the field of legume proteomics to comprehend the defence and regulatory mechanisms during development and adaptation to climatic changes. Here, we attempted to provide a concise overview of the progress in legume proteomics and discuss future developments in three broad perspectives: (i) proteome of organs/tissues; (ii) subcellular compartments; and (iii) spatiotemporal changes in response to stress. Such data mining may aid in discovering potential biomarkers for plant growth, in general, apart from essential components involved in stress tolerance. The prospect of integrating proteome data with genome information from legumes will provide exciting opportunities for plant biologists to achieve long-term goals of crop improvement and sustainable agriculture.