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Induction of hairy root cultures from Gossypium hirsutum and Gossypium barbadense to produce gossypol and related compounds

  • Metabolic Engineering/Secondary Metabolism
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Abstract

Hairy root cultures were induced by inoculating cotyledonary leaves and hypocotyl segments from two cotton species, Gossypium hirsutum and Gossypium barbadense, with Rhizobium rhizogenes 15834. For both species, more hairy roots formed on inoculation sites on cotyledonary leaves than on hypocotyls. The addition of sucrose to basal Murashige–Skoog media increased the frequency of hairy root formation, whereas the addition of naphthalene acetic acid (0.54 μM) did not. After transfer to a liquid culture, hairy root growth was very rapid. After 3 wk in liquid culture, both cotton species produced gossypol, a di-sesquiterpene secondary metabolite with known anticancer activity, and two related methylated derivatives. Most (60–95%) gossypol produced by cultures was retained within the hairy root tissues, but some was found in the media. The average gossypol level observed among 96 different cultures was 15 mg/g of dry culture mass; however, some cultures produced >40 mg/g of dry culture mass. Variation in gossypol levels was greater for cultures arising from different transformation events than for multiple subclones of a single transformant. The high level of gossypol production attained by most of these cultures suggests that they will be valuable for studying the biochemical and molecular aspects of gossypol biosynthesis, capable of producing large amounts of gossypol and related compounds, and useful for generating modified forms of gossypol (e.g., radio-labeled gossypol) for understanding bioactivity mechanisms.

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Abbreviations

HRIM:

hairy root induction media

TE:

transformation event

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Acknowledgments

The authors thank Scott Pelitire for conducting the HPLC analyses and the Louisiana Board of Regents, Governor’s Biotechnology Initiative for providing a research fellowship to Stephanie Moss.

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Authors and Affiliations

  1. Southern Regional Research Center, USDA-ARS, 1100 Robert E. Lee Blvd., New Orleans, LA, 70124, USA

    Barbara A. Triplett, John M. Bland & Michael K. Dowd

  2. Department of Biological Sciences, University of New Orleans, Lakefront, New Orleans, LA, 70148, USA

    Stephanie C. Moss

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  1. Barbara A. Triplett
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  2. Stephanie C. Moss
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  3. John M. Bland
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Correspondence to Barbara A. Triplett.

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Editor: Gregory C. Phillips

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Triplett, B.A., Moss, S.C., Bland, J.M. et al. Induction of hairy root cultures from Gossypium hirsutum and Gossypium barbadense to produce gossypol and related compounds. In Vitro Cell.Dev.Biol.-Plant 44, 508–517 (2008). https://doi.org/10.1007/s11627-008-9141-2

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