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Augmentation of wedelolactone through in vitro tetraploid induction in Eclipta alba (L.) Hassk.

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Abstract

A complete protocol for the in vitro induction of Eclipta alba tetraploids has been optimized to enhance the wedelolactone content, an anti-cancerous compound. The effects of different concentrations of colchicine (0, 0.01, 0.05, 0.1, 0.2 and 0.3%; w/v) along with treatment durations (12, 24, 36 and 48 h) were investigated on shoot tip (ST) and nodal segment (NS). The treated explants were then incubated on Murashige and Skoog (MS) medium having 1.5 mg L−1 N6-benzylaminopurine and 0.5 mg L−1 α-napthalene acetic acid for shoot regeneration and afterward root was induced on 1.0 mg L−1 indole-3-acetic acid enriched ½MS medium. The tetraploids of E. alba were proficiently induced by the treatment of 0.1% colchicine for 24 h. The highest tetraploid induction efficiency was obtained from ST (30.56%) in comparison to the NS (22.22%). Analysis by spectrophotometry and flow cytometry showed that colchicine treated plants contained higher quantity of DNA than diploid plants. Cytological studies demonstrated doubled the chromosome number in tetraploids (2n = 4x = 44) than diploids (2n = 2x = 22). The ploidy level enhancement lead to alteration of other traits, like increased plant height, stem diameter, leaf size, stomatal size and chlorophyll content. As determined through high performance thin-layer chromatography, the ultimate achievement of this technique is the higher accumulation of wedelolactone in tetraploid plants (300.32 µg g−1 dry weight) in evaluation to in vitro diploid (131.31 µg g−1 dry weight) and in vivo diploid mother plants (93.26 µg g−1 dry weight), thus improving the pharmaceutical value of E. alba.

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Abbreviations

BAP:

N6-benzylaminopurine

FCM:

Flow cytometry

HPTLC:

High performance thin-layer chromatography

IAA:

Indole-3-acetic acid

MS:

Murashige and Skoog (1962)

NAA:

α-Napthalene acetic acid

NS:

Nodal segment

RT:

Root tip

ST:

Shoot tip

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Acknowledgements

Authors acknowledge the laboratory as well as library assistance from the Bidhan Chandra Krishi Viswavidyalaya, West Bengal, India.

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    Authors and Affiliations

    1. Department of Agricultural Biotechnology, Faculty of Agriculture, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur, Nadia, West Bengal, 741252, India

      Umme Salma, Suprabuddha Kundu, Md. Nasim Ali & Nirmal Mandal

    2. Faculty Centre for Integrated Rural Development and Management, Ramakrishna Mission Vivekananda University, Ramakrishna Mission Ashrama, Narendrapur, Kolkata, 700103, India

      Alok Kumar Hazra

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    1. Umme Salma
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    2. Suprabuddha Kundu
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    3. Alok Kumar Hazra
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    Contributions

    US, SK, MNA and NM conceived the idea; US and SK performed the experiments; US and SK surveyed the literature and wrote the manuscript; AKH assisted in the phytochemical assessment; MNA and NM edited the manuscript. All the authors approved the final version of the manuscript prior to submission.

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    Correspondence to Umme Salma.

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    The authors declare that they have no conflicts of interest.

    Additional information

    Communicated by Sergio J. Ochatt.

    Umme Salma and Suprabuddha Kundu have contributed equally.

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    Salma, U., Kundu, S., Hazra, A.K. et al. Augmentation of wedelolactone through in vitro tetraploid induction in Eclipta alba (L.) Hassk.. Plant Cell Tiss Organ Cult 133, 289–298 (2018). https://doi.org/10.1007/s11240-018-1381-1

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