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High frequency somatic embryogenesis and synthetic seed production in Clitoria ternatea Linn

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Abstract

An efficient plant regeneration protocol has been developed from embryogenic callus derived from cotyledonary explants of Clitoria ternatea Linn., an important medicinal climber species. Optimum embryogenic callus (75 %) was induced on Murashige and Skoog (MS) medium supplemented with 2 mg/l 2, 4-dichlorophenoxyacetic acid (2, 4-D). On subculturing the callus on MS medium supplemented with 2 mg/l 6-benzyladenine (BA) and 0.5 mg/l α-naphthalene acetic acid (NAA), 61 % of cultures responded with a mean number of 22 somatic embryos per gram callus at different stages of development after 45 days of culture. The addition of higher concentrations of sucrose and abscisic acid (ABA) significantly increased the embryogenic response. MS medium supplemented with 2 mg/l BA, 0.2 mg/l NAA and 4 % of sucrose resulted in 76 % of cultures responding with a mean number of 28 embryos per one gram callus. The highest embryogenic response, frequency of 83 % and mean number of 37 embryos per gram callus, was observed when the MS medium was supplemented with 2 mg/l BA, 0.2 mg/l NAA, and 3 mg/l ABA. Synthetic seeds were produced by encapsulating embryos in calcium alginate gel. The gel contained MS medium with 3 % of sucrose, 1.0 mg/l BA and 0.2 mg/l NAA. The synthetic seeds germinated on MS medium supplemented with BA (1.0-4.0 mg/l) alone or in combination with NAA (0.1–0.7 mg/l) or indole-3-butyric acid (IBA; 0.1–0.7 mg/l). The highest synthetic seed germination (92  %) was observed on MS medium supplemented with 2 mg/l BA and 0.5 mg/l NAA. The synthetic seeds were stored at 4 °C and lab conditions (25 ± 2 °C) up to 5 months. The synthetic seeds kept at 4 °C showed 86 % viability even after 5 months of storage. Both somatic embryos and synthetic seeds germinated and were transferred to soil successfully.

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Abbreviations

2, 4-D:

2, 4-Dichlorophenoxyacetic acid

ABA:

Abscisic acid

BA:

6-benzyladenine

Kn:

Kinetin

MS:

Murashige and Skoog

NAA:

α-naphthalene acetic acid

IBA:

Indole-3-butyric acid

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Acknowledgments

We thank the Principal for providing with necessary laboratory facilities. TDT acknowledges the financial assistance from UGC, Government of India, in the form of a major research project (Project no. 38-233/2009).

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  1. Postgraduate and Research Department of Botany, St. Thomas College, Pala, Arunapuram (P.O.), Kottayam (Dt.), 686574, Kerala, India

    G. Krishna Kumar & T. Dennis Thomas

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  1. G. Krishna Kumar
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Correspondence to T. Dennis Thomas.

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Krishna Kumar, G., Thomas, T.D. High frequency somatic embryogenesis and synthetic seed production in Clitoria ternatea Linn. Plant Cell Tiss Organ Cult 110, 141–151 (2012). https://doi.org/10.1007/s11240-012-0138-5

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