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In vitro regeneration of Melastoma malabatricum Linn. through organogenesis and assessment of clonal and biochemical fidelity using RAPD and HPLC

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Abstract

Melastoma malabatricum Linn. is an important medicinal plant used in folk medicine for the treatment of various infectious diseases. We developed an in vitro protocol for direct adventitious shoot regeneration from leaf explants of M. malabatricum. Murashige and Skoog (MS) was the most efficient basal medium for shoot regeneration. Multiple adventitious shoot formation was higher in medium supplemented with 3 % sucrose than maltose or fructose. Leaf explants cultured on MS medium supplemented with α-naphthaleneacetic acid (NAA) and thidiazuron (TDZ) showed the highest shoot regeneration (78.00 ± 0.58 %) and the largest number of shoots per explant (11.67 ± 3.05). MS supplemented with gibberellic acid (GA3) was the most effective for shoot elongation. The highest number of roots per explant (10.67 ± 3.51) occurred on MS supplemented with indole-3-butyric acid (IBA), indicating the IBA was more effective to induce rooting than indole acetic acid. Explants with intact petioles and lamina were more responsive and produced a larger number of shoots per explant (14.67 ± 2.52) than explants with lamina alone (8.00 ± 2.00). Histology and scanning electron microscopy of regenerated shoots confirmed the occurrence of direct organogenesis. Random amplified polymorphic DNA analysis confirmed that in vitro regenerated plants were genetically similar to their mother plant. High-performance liquid chromatography of phenolic acids in leaf extracts of regenerants revealed no significant differences in the phenolic compound profile compared with mother plants. Our in vitro regeneration protocol represents a valuable tool for germplasm conservation and genetic transformation of M. malabatricum.

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Abbreviations

PGR:

Plant growth regulator

NAA:

Naphthalene acetic acid

IBA:

Indole-3-butyric acid

BA:

6-Benzyladenine

TDZ:

Thidiazuron

IAA:

Indole acetic acid

MS:

Murashige and Skoog

SH:

Schenk and Hildebrandt

B5:

Gamborg et al. medium

SEM:

Scanning electron microscopy

GA3 :

Gibberellic acid

FAA:

Formalin–acetic acid–alcohol

RAPD:

Random amplified polymorphic DNA

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Acknowledgments

This research was supported by the 2015 KU Brain Pool of Konkuk University.

Author contributions

Bimal Kumar Ghimire performed experiments for initial regeneration method development. Eun Soo Seong performed DNA isolation and RAPD analysis. Truong Xuan Nguyen performed assessment of the ploidy level of regenerated plants. Chang Yeon Yu and Seung Hyun Kim provided technical support and revised the manuscript critically. Ill-Min Chung performed phytochemical analysis of regenerants, revised the manuscript and supervised the study as a principal investigator. All authors read and approved the final manuscript.

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Authors and Affiliations

  1. Department of Applied Life Science, Konkuk University, Seoul, 143-701, South Korea

    Bimal Kumar Ghimire, Seung Hyun Kim & Ill-Min Chung

  2. Bioherb Research Institute, Kangwon National University, Chuncheon, 200-701, South Korea

    Eun Soo Seong & Chang Yeon Yu

  3. Department of Horticulture, Kangwon National University, Chuncheon, 200-701, South Korea

    Truong Xuan Nguyen

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  1. Bimal Kumar Ghimire
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  2. Eun Soo Seong
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  3. Truong Xuan Nguyen
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Correspondence to Ill-Min Chung.

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Ghimire, B.K., Seong, E.S., Nguyen, T.X. et al. In vitro regeneration of Melastoma malabatricum Linn. through organogenesis and assessment of clonal and biochemical fidelity using RAPD and HPLC. Plant Cell Tiss Organ Cult 124, 517–529 (2016). https://doi.org/10.1007/s11240-015-0911-3

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