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Direct shoot regeneration from nodal, internodal and petiolar segments of Piper longum L. and in vitro conservation of indexed plantlets

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Abstract

Three explants namely, nodal, internodal and petiolar segments were used to establish in vitro cultures of Piper longum. Multiple shoots were induced on semi-solid Murashige and Skoog (MS) medium supplemented with 1 mg/l 6-benzyladenine (BA). Addition of ascorbic acid (40 mg/l) considerably reduced browning of tissue and medium. Best shoot regeneration was observed from petiolar explants and was, therefore, used for all further studies. An indexing method was introduced for checking bacterial contamination in well established shoot multiplication cultures. It was found that bacterial infection was quite high in shoots derived from nodal and internodal explants while it was least in those obtained from petiolar segments. Only shoots that indexed negative for endogenous bacteria were used for proliferation and in vitro conservation studies. At the end of 4 weeks in proliferation medium which consisted of MS supplemented with 0.5 mg/l BA and 40 mg/l ascorbic acid as many as 22 shoot buds of 41 mm length could be obtained. Shoot buds developed into clusters for ease of further proliferation. A step of shoot elongation for 2 weeks in liquid MS basal medium was found to be beneficial for getting long and healthy shoots for rooting. Single shoots were rooted in 0.25 mg/l indole butyric acid that could be successfully acclimatized under nethouse conditions. A conservation strategy was also developed. The shoot cultures could be maintained without subculturing for as long as 8 weeks in MS medium supplemented with 1 mg/l paclobutrazol (PBZ) and 40 mg/l ascorbic acid.

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Abbreviations

BA:

6-Benzyladenine

IBA:

Indole butyric acid

Kn:

Kinetin

MS:

Murashige and Skoog

PBZ:

Paclobutrazol

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Acknowledgments

PKD gratefully acknowledges the University Grants Commission, New Delhi, for the financial help in the form of a Major Project vide letter no. F.32-382/2006 (SR). The authors thank the Director of the Institute for providing the necessary laboratory facilities. National Bureau of Plant Genetic Resources, Regional Station, Thrisshur, Kerela, is gratefully acknowledged for the supply of Piper longum plant material. Professor Sant S. Bhojwani is thankfully acknowledged for going through the manuscript.

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  1. Plant Biotechnology Laboratory, Department of Botany, Dayalbagh Educational Institute (Deemed University), Dayalbagh, Agra, 282110, India

    Dolly Rani & Prem Kumar Dantu

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  1. Dolly Rani
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  2. Prem Kumar Dantu
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Correspondence to Prem Kumar Dantu.

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Rani, D., Dantu, P.K. Direct shoot regeneration from nodal, internodal and petiolar segments of Piper longum L. and in vitro conservation of indexed plantlets. Plant Cell Tiss Organ Cult 109, 9–17 (2012). https://doi.org/10.1007/s11240-011-0068-7

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