Abstract
An efficient regeneration protocol for the medicinal plant, Paris polyphylla Sm. was developed through the formation of mini-rhizomes (MRs) using transverse thin cell layer (tTCL) culture technique. MRs were induced from tTCL explants derived from the basal and middle stem portions while apical portion failed to show any kind of response. Highest response percentage (86.6 %) of MRs formation with a maximum fresh weight (1.05 ± 0.08 g) was achieved from basal sections cultured on ½ MS medium supplemented with 0.5 mg/l 6-benzylaminopurine (BAP). MRs transferred to plant growth regulator free medium gave rise to shoot buds that eventually regenerated into plantlets and were successfully acclimatized with a survival percentage of more than 95 % under greenhouse conditions. Quantification through reverse-phase HPLC showed 1.41-fold higher content of total steroidal saponins in MRs cultured on medium supplemented with 0.5 mg/l BAP as compared to the field-grown rhizome. Elicitation of MRs liquid culture with chitosan, salicyclic acid (SA) and yeast extract enhanced the production of steroidal saponins but resulted in reduced growth rate. Highest total steroidal saponins content (87.66 ± 1.66 mg/g DW) was achieved in cultures treated with SA at 50 mg/l after 30 days of elicitation which is 3.6 times higher than the in vivo rhizome. The developed protocol would facilitate the conservation of this valuable medicinal plant and could be used as a ready stock to meet the demands of the pharmaceutical industry for steroidal saponins productions.
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Abbreviations
- BAP:
-
6-Benzylaminopurine
- CHI:
-
Chitosan
- DW:
-
Dry weight
- FW:
-
Fresh weight
- HPLC:
-
High performance liquid chromatography
- KIN:
-
Kinetin
- MR:
-
Mini-rhizome
- MS:
-
Murashige and Skoog
- SA:
-
Salicyclic acid
- TDZ:
-
Thidiazuron
- tTCL:
-
Transverse thin cell layer
- PGR:
-
Plant growth regulator
- YE:
-
Yeast extract
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Acknowledgments
The authors acknowledge Dr. A. Bhattacharjee and Ms. B. J. Mylliemngap, Department of Biotechnology and Bioinformatics, North-Eastern Hill University, Shillong for providing the HPLC facilities and valuable help. The authors would also like to thank Prof. N. Venugopal, Department of Botany, North-Eastern Hill University, Shillong for permission to use microtome. SR is thankful to University Grant Commission, India for awarding her Rajiv Gandhi National Fellowship for SC/ST.
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Raomai, S., Kumaria, S., Kehie, M. et al. Plantlet regeneration of Paris polyphylla Sm. via thin cell layer culture and enhancement of steroidal saponins in mini-rhizome cultures using elicitors. Plant Growth Regul 75, 341–353 (2015). https://doi.org/10.1007/s10725-014-9957-1
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DOI: https://doi.org/10.1007/s10725-014-9957-1