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<p>An effective micropropagation method for the valuable medicinal plant <i>Hydrocotyle sibthorpioides</i> has been developed. Aseptic nodal segments were inoculated onto Murashige and Skoog (MS) medium augmented with 0.6&#xa0;µM 6-benzyladenine (BA), which was found to be effective for multiple shoot induction, with a mean shoot number of 10.22 shoots per culture. However, continuous subculture on a 6-benzyladenine added medium caused shoot quality deterioration. Therefore, at the third subculture onwards, the growth retardant, daminozide (DAM), was supplemented to the Murashige and Skoog medium at varying concentrations to compare its effect on multiple shoot production with 6-benzyladenine. A significantly higher number of multiple shoots (16.22) of better quality was produced on 5.0&#xa0;µM daminozide, including 1.5-fold higher than medium supplemented with 0.6&#xa0;µM 6-benzyladenine. Cultures grown on 5.0&#xa0;µM daminozide showed a good number of roots, which is helpful in the acclimatization of the plantlets without auxin treatments. Using inter simple sequence repeat (ISSR) and start codon targeted (SCoT) markers, the genetic uniformity of the regenerated plants was validated, where a higher degree of monomorphism with the mother plant was recorded. The present study indicated that substituting 6-benzyladenine with daminozide would promote multiple shoot production, coupled with a high rate of survival of microshoots during rooting and hardening. In routine micropropagation, continuous use of cytokinins like 6-benzyladenine often causes excessive shoot elongation and hyperhydricity, leading to poor quality plants. The present findings suggested that replacing 6-benzyladenine with daminozide in the subculture medium significantly improved shoot quality, controlled elongation, reduced hyperhydricity, and better survival during acclimatization. The daminozide-based micropropagation system, therefore, can be extended to other plant species that struggle with poor shoot quality and abnormal internodal elongation, especially in the case of economically important crop species.</p>

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Growth retardant daminozide effectively induces microshoots in cytokinin-devoid in vitro cultures of Hydrocotyle sibthorpioides Lam.

  • 期刊论文

  • 发表日期：

    2026年3月17日

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  • James Silpa,
  • Bindu R. Nair,
  • E. A. Siril

摘要

An effective micropropagation method for the valuable medicinal plant Hydrocotyle sibthorpioides has been developed. Aseptic nodal segments were inoculated onto Murashige and Skoog (MS) medium augmented with 0.6 µM 6-benzyladenine (BA), which was found to be effective for multiple shoot induction, with a mean shoot number of 10.22 shoots per culture. However, continuous subculture on a 6-benzyladenine added medium caused shoot quality deterioration. Therefore, at the third subculture onwards, the growth retardant, daminozide (DAM), was supplemented to the Murashige and Skoog medium at varying concentrations to compare its effect on multiple shoot production with 6-benzyladenine. A significantly higher number of multiple shoots (16.22) of better quality was produced on 5.0 µM daminozide, including 1.5-fold higher than medium supplemented with 0.6 µM 6-benzyladenine. Cultures grown on 5.0 µM daminozide showed a good number of roots, which is helpful in the acclimatization of the plantlets without auxin treatments. Using inter simple sequence repeat (ISSR) and start codon targeted (SCoT) markers, the genetic uniformity of the regenerated plants was validated, where a higher degree of monomorphism with the mother plant was recorded. The present study indicated that substituting 6-benzyladenine with daminozide would promote multiple shoot production, coupled with a high rate of survival of microshoots during rooting and hardening. In routine micropropagation, continuous use of cytokinins like 6-benzyladenine often causes excessive shoot elongation and hyperhydricity, leading to poor quality plants. The present findings suggested that replacing 6-benzyladenine with daminozide in the subculture medium significantly improved shoot quality, controlled elongation, reduced hyperhydricity, and better survival during acclimatization. The daminozide-based micropropagation system, therefore, can be extended to other plant species that struggle with poor shoot quality and abnormal internodal elongation, especially in the case of economically important crop species.