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Mass propagation through direct and indirect organogenesis in three species of genus Zephyranthes and ploidy assessment of regenerants through flow cytometry

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Abstract

Genus Zephyranthes consists of economically important plant species due to their high ornamental value and presence of valuable bioactive compounds. However, this genus propagates by asexual division only which gives slow propagation rate. Plant tissue culture has the potential to provide efficient techniques for rapid multiplication and genetic improvement of the genus. In this work, a dual in vitro regeneration system through callus mediated shoot regeneration and direct shoot regeneration in species Zephyranthes candida, Zephyranthes grandiflora and Zephyranthes citrina was investigated. Bulb, leaf and root explants were cultured on Murashige and Skoog (MS) medium amended with different plant growth regulators (PGR’s) viz. 2,4-dichlorophenoxyacetic acid (2,4-D), 1-Naphthalene acetic acid (NAA), 6-benzyl amino purine (BAP), N-phenyl-N′-1,2,3 -thiadiazol-5-ylurea (TDZ), 6-Furfuryl- aminopurine (KIN) alone or in combinations for callus induction and regeneration. Only bulb explants showed callus induction and regeneration response on different PGR combinations with a varied response in callus induction percentage, callus color and callus texture. Creamish compact callus (CC) was induced on 2 mg L\(^{-1}\) 2,4-D, brown friable callus (BF) on 2 mg L\(^{-1}\) NAA + 1 mg L\(^{-1}\) BAP and green friable callus (GF) callus on 1 mg L\(^{-1}\) KIN + 3 mg L\(^{-1}\) NAA. The maximum shoot multiplication from different callus types (indirect organogenesis) was achieved on 2 mg L\(^{-1}\) BAP alone without combinations. Bulb explants of Z. grandiflora induced maximum callus induction percentage (86.4%) and shoot regeneration percentage (83.5%) with the maximum 08 shoots per 150 mg callus mass. The induction and regeneration response was followed in the order of Z. grandiflora > Z. candida > Z. citrina. Similarly, maximum direct organogenesis from bulb explants was obtained in Z. grandiflora (93.3%) followed by Z. candida (91.5%) and Z. citrina (90.4%) on 3 mg L\(^{-1}\) TDZ amended MS media. Adventitious root induction was achieved on 2 mg L\(^{-1}\) IBA with a maximum of 8 roots per shoot. The in vitro raised plantlets were successfully acclimatized in the field with 85% survival efficiency. The genome size (2C DNA content) of the field-grown plants and in vitro regenerated plants, evaluated through flow cytometry technique, were similar and showed no ploidy changes. An efficient mass propagation protocol was established for obtaining plants with unaltered genome size in the three species of Zephyranthes.

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Abbreviations

MS:

Murashige and Skoog medium

PGRs:

Plant growth regulators

2,4-D:

2,4-Dichlorophenoxyacetic acid

NAA:

Naphthalene acetic acid

IBA:

Indole-3-butyric acid

BAP:

6-Benzylaminopurine

TDZ:

Thidiazuron

KIN:

Kinetin

pg:

Picograms

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Acknowledgements

Authors are thankful to the Department of Botany, Jamia Hamdard for providing laboratory facilities, Dr. Shwetanjali Nimker (Application Scientist) at BD-FACS, Jamia Hamdard, New Delhi, for providing flow cytometry facility; Dr Akhtar H Malik (Center for Biodiversity and Taxonomy, Department of Botany, University of Kashmir, Srinagar, India) for plant identification.

Funding

The doctoral research funding from Hamdard National Foundation, New Delhi, India in favor of the first author is duly acknowledged.

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Authors and Affiliations

  1. Cellular Differentiation and Molecular Genetics Section, Department of Botany, Jamia Hamdard, New Delhi, India

    Rukaya Syeed, A. Mujib, Moien Qadir Malik, Jyoti Mamgain, Bushra Ejaz, Basit Gulzar & Nadia Zafar

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  1. Rukaya Syeed
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  2. A. Mujib
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Contributions

RS performed all the experiments and made the manuscript; others helped in making tables, figures and photoplates; and AM edited the manuscript before submission.

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Correspondence to A. Mujib.

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Syeed, R., Mujib, A., Malik, M.Q. et al. Mass propagation through direct and indirect organogenesis in three species of genus Zephyranthes and ploidy assessment of regenerants through flow cytometry. Mol Biol Rep 48, 513–526 (2021). https://doi.org/10.1007/s11033-020-06083-1

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