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Bergenia ligulata (Wall.): micropropagation, genetic fidelity, and SEM studies

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Abstract

Bergenia ligulata, commonly known as ‘Pashanbheda’ or Indian rhubarb, is a perennial herb that has been recognized for its diverse medicinal properties. The indiscriminate use of B. ligulata has brought the species to the brink of becoming threatened. This research aims to establish a robust tissue culture protocol that can be utilized for the rapid micropropagation of B. ligulata. This protocol is essential for ensuring the sustainable production of this valuable plant species and preventing the depletion of its natural populations. The study successfully demonstrated an efficient in vitro regeneration in B. ligulata, using leaf and petiole explants. The most effective combination for achieving the highest number of shoots on either explant (leaf or petiole) involved using Murashige and Skoog (MS) medium supplemented with 0.9 µM and 1.8 µM 6-benzylamino purine (BAP) with 0.5 µM 1-naphthaleneacetic acid (NAA). Moreover, multiple shoots were also produced on MS medium fortified with 8.8 µM BAP and 2.3 µM kinetin (Kn). To achieve optimal rooting, the 45-d-old shoot was carefully isolated and placed in a half-strength MS medium. PCR-based molecular analysis using inter simple sequence repeats (ISSR) confirmed the genetically clonal nature of regenerated plantlets. About 80% of the well-developed in vitro regenerated plants were acclimatized in the glasshouse, thereby showing the robustness of the developed protocol. Based on the present study, a reproducible in vitro technique was utilized to achieve direct regeneration of approximately 3597 plants from a single explant over a 1-yr period. This approach involved molecular fidelity analysis and scanning electron microscopy (SEM) analyses to ensure reliable results.

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References

  • Akter KT, Hoque MdA (2015) Effect of IBA and BAP on in vitro plant regeneration of mint. J Crop Sci and Tech 4:1–10

    Google Scholar 

  • Amiri S, Mohammadi R (2021) Establishment of an efficient in vitro propagation protocol for Sumac (Rhus coriaria L.) and confirmation of the genetic homogeneity. Sci Rep 11:173

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ashraf MF, Aziz MA, Kemet II (2014) Effect of cytokinin types, concentrations and their interactions on in vitro shoot regeneration of Chlorophytum borivilianum Sant. & Fernandez. Elec J Biotechnol 17:275–279

    Article  Google Scholar 

  • Balilashaki K, Vahedi M, Karimi R (2015) In vitro direct regeneration from node and leaf explants of Phalaenopsis cv. ‘Surabaya. Plant Tiss Cult Biotech 25:193–205

    Article  Google Scholar 

  • Bashir S, Gilani AH (2009) Antiurolithic effect of Bergenia ligulata rhizome: an explanation of the underlying mechanisms. J Ethnopharmacol 122:106–116

    Article  PubMed  Google Scholar 

  • Bhandari MR, Jong-Anurakkun N, Hong G, Kawabata J (2008) α-Glucosidase and α-amylase inhibitory activities of Nepalese medicinal herb Pakhanbhed (Bergenia ciliata, Haw.). Food Chem 106:247–252

    Article  CAS  Google Scholar 

  • Bhatt R, Arif M, Gaur MK, Rao PB (2008) Rauwolfia serpentina: protocol optimization for in vitro propagation. African J Biotechnol 7:4265–4268

    CAS  Google Scholar 

  • Dalila ZD, Jaafar H, Manaf AA (2013) Effects of 2,4-D and kinetin on callus induction of Barringtonia racemosa leaf and endosperm explants in different types of basal media. Asian J Plant Sci 12:21–27

    Article  CAS  Google Scholar 

  • Dhar ML, Dhar MM, Dhawan BN, Mehrotra BN, Ray C (1968) Screening of Indian plants for biological activity: part I. Indian J Exp Biol 6:232–247

    CAS  PubMed  Google Scholar 

  • Garimella TS, Jolly CI, Narayanan S (2001) In vitro studies on antilithiatic activity of seeds of Dolichos biflorus Linn. and rhizomes of Bergenia ligulata Wall. Phytother Res 15:351–355

    Article  CAS  PubMed  Google Scholar 

  • Gurav SS, Gurav NS (2014) A comprehensive review: Bergenia ligulata Wall- a controversial clinical candidate. Int J Pharm Sci Res 5:1630

    Google Scholar 

  • Goel AK, Kulshreshtha DK, Dubey MP, Rajendran SM (2002) Screening of Indian plants for biological activity: Part XVI. Indian J Exp Biol 40:812–827

    CAS  PubMed  Google Scholar 

  • Gohain A, Sharma A, Gogoi HJ, Cooper R, Kaur R, Nayik GA, Shaikh AM, Kovács B, Areche FO, Ansari MJ, Alabdallah NM, Al-Farga A (2022) Bergenia pacumbis (Buch.-Ham. ex D.Don) C.Y.Wu & J.T.Pan: a comprehensive review on traditional uses. Plants (Basel) 11:1129

    Article  CAS  PubMed  Google Scholar 

  • Grzegorczyk-Karolak I, Kuźma Ł, Wysokińska H (2016) In vitro cultures of Scutellaria alpina as a source of pharmacologically active metabolites. Acta Physiol Plant 38:1–9

    Article  CAS  Google Scholar 

  • Gürocak S, Küpeli B (2006) Consumption of historical and current phytotherapeutic agents for urolithiasis: a critical review. J Urol 176:450–455

    Article  PubMed  Google Scholar 

  • Haq I, Khurshid G, Abbasi AZ, Nawaz I, Naqvi T, Arfan M, Ali MA, Rehman S (2019) Successful callogenesis from leaf and petiole of Bergenia ciliata (Haw) Sternb and antibacterial activity of callus extracts. Pak J Bot 51:1847–1852. https://doi.org/10.30848/PJB2019-5(11)

    Article  CAS  Google Scholar 

  • Kashima Y, Yamaki H, Suzuki T, Miyazawa M (2011) Insecticidal effect and chemical composition of the volatile oil from Bergenia ligulata. J Agric Food Chem 13:7114–71149

    Article  Google Scholar 

  • Khurana-Kaul V, Kachhwaha S, Kothari SL (2010) Direct shoot regeneration from leaf explants of Jatropha curcas in response to thidiazuron and high copper contents in the medium. Biol Plant 54:369–372

    Article  CAS  Google Scholar 

  • Mok DWS, Mok MC (2001) Cytokinin metabolism, and action. Annu Rev Plant Physiol Plant Mol Biol 52:89–118

    Article  CAS  PubMed  Google Scholar 

  • Moreau RA, Whitaker BD, Hicks KB (2002) Phytosterols, phytosterols, and their conjugates in foods: structural diversity, quantitative analysis, and health-promoting uses. Prog Lipid Res 41:457–500. https://doi.org/10.1016/s0163-7827(02)00006-1

    Article  CAS  PubMed  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiol Plant 15:473–497. https://doi.org/10.1111/j.1399-3054.1962.tb08052.x

    Article  CAS  Google Scholar 

  • Nasri F, Mortazavi NS, Ghaderi N, Javadi T (2013) Propagation in vitro of Alstroemeria ligtu hybrid through direct organogenesis from the leaf base. J Hort Res 21:23–30

    CAS  Google Scholar 

  • Nasir Nor AN, Md AN, Zainuddin Z, Yaakob Z (2017) Evaluation of genetic homogeneity of Jatropha curcas L. hybrid at an early stage of shoot bud formation from petioles using ISSR marker. Biotechnol Biotechnol Equip 31:45–50. https://doi.org/10.1080/13102818.2016.1257923

    Article  CAS  Google Scholar 

  • Oberhofer M, Malfent F, Zehl M, Urban E, Wackerlig J, Reznicek G, Vignolle GA, Rückert C, Busche T, Wibberg D, Zotchev SB (2022) Biosynthetic potential of the endophytic fungus Helotiales sp. BL73 revealed via compound identification and genome mining. Appl Environ Microbiol 88:e0251021. https://doi.org/10.1128/aem.02510-21

    Article  PubMed  Google Scholar 

  • Pandey A, Chandra Sekar K, Tamta S, Rawal RS (2018) Assessment of phytochemicals, antioxidant and antimutagenic activity in micro-propagated plants of Quercus serrata, a high-value tree species of Himalaya. Plant Biosyst 152:929–936. https://doi.org/10.1080/11263504.2017.1395372

    Article  Google Scholar 

  • Plummer J (2020) Bergenia pacumbis. The IUCN red list of threatened species 2020:e.T156235116A156238222.https://doi.org/10.2305/IUCN.UK.20203.RLTS.T156235116A156238222.en

  • Rafi S, Kamili AN, Ganai BA, Mir MY, Parray JA (2016) In vitro culture and biochemical attributes of Bergenia ciliata (Haw.) Sternb. Proc Natl Acad Sci India Sect B Biol Sci 88:609–619. https://doi.org/10.1007/s40011-016-0797-9

    Article  CAS  Google Scholar 

  • Rajbhandari M, Mentel R, Jha PK, Chaudhary RP, Bhattarai S, Gewali MB, Karmacharya N, Hipper M, Lindequist U (2009) Antiviral activity of some plants used in Nepalese traditional medicine. Evid Based Complement Alternat Med 1:517–522

    Article  Google Scholar 

  • Rajbhandari M, Wegner U, Jülich M, Schoepke T, Mentel R (2001) Screening of Nepalese medicinal plants for antiviral activity. J Ethnopharmacol 74:251–255

    Article  CAS  PubMed  Google Scholar 

  • Rasool R, Kamili AN, Ganai BA, Akbar S (2009) Effect of BAP and NAA on shoot regeneration in Prunella vulgaris. J Nat Sci and Math 3:21–26

    Google Scholar 

  • Ruby KM, Dwivedi J, Chauhan R (2012) Pashanbheda a golden herb of Himalaya: a review. Int J Pharm Rev Res 2:97–105

    Google Scholar 

  • Sahoo S, Rout GR (2014) Plant regeneration from leaf explants of Aloe barbadensis Mill. and genetic fidelity assessment through DNA markers. Physiol Mol Biol Plants 20:235–240. https://doi.org/10.1007/s12298-014-0226-6

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Saijyo J, Suzuki Y, Okuno Y, Yamaki H, Suzuki T, Miyazawa M (2008) Alpha-glucosidase inhibitor from Bergenia ligulata. J Oleo Sci 57:431–435. https://doi.org/10.5650/jos.57.431

    Article  CAS  PubMed  Google Scholar 

  • Salehi M, Hosseini B, Jabbarzadeh Z (2014) High-frequency in vitro plantlet regeneration from apical bud as a novel explant of Carum copticum L. Asian Pac J Trop Biomed 4:424–428

    Article  Google Scholar 

  • Sharma HK, Chhangte L, Dolui AK (2001) Traditional medicinal plants in Mizoram, India. Fitoterapia 72:146–161

    Article  CAS  PubMed  Google Scholar 

  • Sharma N, Chandel KPS, Anderson P (1993) In vitro propagation of Gentiana kurroo — an indigenous threatened plant of medicinal importance. Plant Cell Tiss Org Cult 34:307–309

    Article  CAS  Google Scholar 

  • Shirin F, Rana PK (2007) In vitro plantlet regeneration from nodal explants of field-grown culms in Bambusa glaucescens Willd. Plant Biotechnol Rep 1:141–147

    Article  Google Scholar 

  • Singh R (2015) Medicinal plants: a review. J Plant Sci 3:50

    Google Scholar 

  • Singh M, Chaturvedi R (2013) Somatic embryogenesis in Neem (Azadirachta indica A. Juss.): current status and biotechnological perspectives. Somatic embryogenesis and Gene expression, Narosa Publishing House, New Delhi, pp 35–55

  • Singh M, Pandey A (2019) In vitro propagation of Bergenia ciliata Sternb: a valuable medicinal and ornamental plant of Sikkim Himalaya. Med Plants 11:117–120

    Google Scholar 

  • Singh N, Juyal V, Gupta A, Gahlot M, Prashant U (2009) Antidiabetic activity of ethanolic extract of the root of Bergenia ligulata in alloxan diabetic rats. Indian Drugs 46:247–249

    Google Scholar 

  • Singh V, Chauhan NS, Singh M, Idris A, Madanala R, Pande V, Mohanty CS (2014) Establishment of an efficient and rapid method of multiple shoot regeneration and a comparative phenolics profile in in vitro and greenhouse-grown plants of Psophocarpus tetragonolobus (L.) DC. Plant Signal Behav 9:e970443. https://doi.org/10.4161/15592316.2014.970443

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Thakur AK, Srivastava DK (2006) High-efficiency plant regeneration from leaf explants of male Himalayan poplar (Populus ciliata Wall.). In Vitro Cell Dev Biol – Plant 42:144–147

    Article  CAS  Google Scholar 

  • Tilkat E, Onay A (2009) Direct shoot organogenesis from in vitro-derived mature leaf explants of pistachio. In Vitro Cell Dev Biol - Plant 45:92–98

    Article  CAS  Google Scholar 

  • Touchell D, Smith J, Ranney TG (2008) Organogenesis from Hypericum frondosum leaves. SNA Research Conference Vol. 53

  • Tuly JA, Hoque MA (2015) In-vitro plantlet regeneration in mint genotypes using different explants. J Crop Sci Technol 4:24–33

    Google Scholar 

  • Yoong LD, Kwame KT, Chaw L (2019) Effects of different combination concentrations of BAP and NAA on types of explants and their regeneration. Int J Agric Sci 9:1–5

    Google Scholar 

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Acknowledgements

The authors are grateful to the Director CSIR-CIMAP for providing basic infrastructure, instrumental facilities, and lab space for conducting the experiments. The authors are also thankful to the Department of Science and Technology, New Delhi, for providing Women Scientist Fellowship-A (SR/WOS-A/LS-226/2018) for the project based on Bergenia ligulata. The manuscript has been assigned a communication number of CIMAP/PUB/2022/126 by CSIR-CIMAP.

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Authors and Affiliations

  1. Biotechnology Department, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow, 226015, India

    Farah Deeba, Zafar Iqbal Warsi, Zakir Husain, Mamta Kumari & Laiq-Ur Rahman

  2. Biological Central Facility: BCF, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow, 226015, India

    Pooja Singh

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  1. Farah Deeba
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Correspondence to Laiq-Ur Rahman.

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Deeba, F., Singh, P., Warsi, Z.I. et al. Bergenia ligulata (Wall.): micropropagation, genetic fidelity, and SEM studies. In Vitro Cell.Dev.Biol.-Plant 60, 75–84 (2024). https://doi.org/10.1007/s11627-023-10398-6

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