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Callus induction and efficient in vitro plant regeneration protocol for Chickpea

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Abstract

The development of an efficient and consistent callus-mediated in vitro regeneration protocol is crucial for biotechnological approaches aimed at improving chickpea, an economically important crop legume. In this study, we assess the effectiveness of callus-mediated regeneration in different chickpea genotypes. Through in vitro screening of explants, we identified the Indian cultivar Pusa 240 as a favourable genotype with higher efficiency of somatic embryogenesis and in vitro plant regeneration. Building upon this finding, we have successfully established two distinct protocols for chickpea callus-mediated somatic embryogenesis, utilizing leaf and hypocotyl explants obtained from the Pusa 240 genotype. These protocols achieved plant regeneration efficiencies of 27% using leaf explants and 46.6 − 66% using hypocotyl explants. Extensive literature review and comparative analysis underscored the superiority of our current protocol. Subsequently, the regenerated plants were successfully acclimatized and transferred to the greenhouse, exhibiting normal phenotypic growth. This detailed regeneration method will provide a valuable resource for chickpea genetic transformations and the generation of large mutant populations where embryogenesis via callus formation is required. The protocol presented here establishes a powerful tool for studying the functional genomics of chickpea plants and lays the foundation for future advancements in this field.

Key message

Identification of the efficient genotype and development of an efficient callus-mediated somatic embryogenesis protocol for chickpea transformation. An essential milestone towards chickpea Tnt1 based mutant population generation.

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Data availability

This manuscript has data included as electronic supplemental file.

Abbreviations

2,4-D:

2,4-Dichlorophenoxyacetic acid

BAP:

6-benzylaminopurine

CIM:

Callus-inducing medium

CK-like:

Cytokinin-like

GA3 :

Gibberellic acid 3

IBA:

Indole-3-butyric acid

MS:

Murashige and Skoog (1962) medium

NAA:

a-naphthalene acetic acid

RIM:

Root-inducing medium

SH:

Schenk-Hildebrandt (1972) medium

SIM:

Shoot-inducing medium

SEM:

Shoot elongation medium

TDZ:

Thidiazuron

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Acknowledgements

The authors would like to thank Dr S.K. Parida, NIPGR New Delhi, for providing chickpea seeds. We acknowledge NIPGR Phytotron Facility; CIF-NIPGR and DBT (Department of Biotechnology)-eLibrary Consortium (DeLCON), India, for providing access to e-resources. We acknowledge the technical assistance of Pradeep Maurya.

Funding

This work was financially supported by research grants from NIPGR core and DBT (BT/PR38393/GET/119/302/2020).

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Authors and Affiliations

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

    Vinod Kumar Jangid, Muthappa Senthil-Kumar & Senjuti Sinharoy

  2. Regional Centre for Biotechnology, New Delhi, India

    Divya Chandran

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  1. Vinod Kumar Jangid
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  2. Muthappa Senthil-Kumar
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  3. Divya Chandran
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Contributions

The conception of the study and experiments were designed by SS, MSK and DC. VKJ performed experiments, analysed data, and prepared the figures. VKJ and SS wrote the manuscript. All authors commented on and contributed to the revision of the manuscript. All authors read and approved the manuscript for publication.

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Correspondence to Senjuti Sinharoy.

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The authors declare that there is no conflict of interest.

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The work was conducted as per the Institutional Biosafety Committee (IBSC) protocol.

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Communicated by Sergio J. Ochatt.

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Jangid, V.K., Senthil-Kumar, M., Chandran, D. et al. Callus induction and efficient in vitro plant regeneration protocol for Chickpea. Plant Cell Tiss Organ Cult 156, 21 (2024). https://doi.org/10.1007/s11240-023-02633-0

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