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Hairy root induction and polysaccharide production of medicinal plant Callerya speciosa Champ.

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Abstract

A reliable and efficient hairy root induction system for the important medicinal plant Callerya speciosa was established by optimizing several parameters that affected the efficiency of Agrobacterium rhizogenes-mediated transformations. Four strains, A4, LBA9402, R1601 and C58C1, and four types of explants, hypocotyls, cotyledons, leaves and excised stems, were examined and the highest rate of transformation (68.3 %) was obtained in cotyledons using strain LBA9402. Therefore, the optimum co-cultivation time and acetosyringone (AS) concentration were evaluated for LBA9402-infected cotyledons and determined to be 2 days and 100 µM AS, respectively, resulting in 64.4 and 66.7 % hairy root transformation efficiency rates, respectively. Transgenic hairy root lines were confirmed by PCR amplification and Southern blot. The growth trend of hairy roots was “slow-quick-slow” and peaked after 30 days of culturing, with increases of 11-fold and 11.4-fold of fresh and dry weights respectively, compared with initial inoculation. Hairy roots showed the ability to synthesize polysaccharides. The effects of different media Murashige and Skoog (MS), ½ MS, WPM and B5, and carbon sources, sucrose, glucose, fructose and maltose on the growth and polysaccharide production of hairy roots were also studied. The ½ MS medium was found to be superior, resulting in the highest accumulation of biomass (10.9 g L−1 fresh weight and 1.1 g L−1 dry weight), the maximum growth index (9.96) and polysaccharide content (22.56 mg g−1 dry weight). Sucrose at 30 g L−1 was optimal for both biomass accumulation and polysaccharide production.

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Abbreviations

6-BA:

6-Benzyladenine

MS:

Murashige and Skoog

NAA:

a-Naphthaleneacetic acid

AS:

Acetosyringone

Cef:

Cefotaxime

YEB:

Yeast extract broth

YMB:

Yeast mannitol broth

PCR:

Polymerase chain reaction

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Acknowledgments

This work was supported by a grant from the Guangxi Natural Science Foundation of China (No. 2013GXNSFBA019173) and Guangxi Science Committee Projects of China (Nos. 14124002-2 and 20133032-5). The authors would like to thank Dr. Dev Sooranna, Imperial College London, for editing the manuscript.

Author's contributions

L. B. designed the experimental procedures and revised the manuscript. S. Y. conducted the induction, identification and growth of hairy roots; the determination of different parameters; the measurement of FW, DW and polysaccharide; and analyzed the data and wrote the manuscript. Z. L. performed the experiments regarding to bacterial strain culturing. Y. Z. and H. H. cultured the plants. R. W. and T. M. performed the PCR amplification and Southern blot. All of the authors read and approved the final manuscript.

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Authors and Affiliations

  1. Guangxi Botanical Garden of Medicinal Plants, Chang-gang Road 189, Nanning, 530023, China

    Shao-chang Yao, Long-hua Bai, Zu-zai Lan, Mei-qiong Tang, Yong-jin Zhai, Hao Huang & Rong-chang Wei

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  1. Shao-chang Yao
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  2. Long-hua Bai
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Correspondence to Shao-chang Yao.

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Yao, Sc., Bai, Lh., Lan, Zz. et al. Hairy root induction and polysaccharide production of medicinal plant Callerya speciosa Champ.. Plant Cell Tiss Organ Cult 126, 177–186 (2016). https://doi.org/10.1007/s11240-016-0988-3

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  • DOI: https://doi.org/10.1007/s11240-016-0988-3

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