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Optimization of methods for using polyethylene glycol as a non-permeating osmoticum for the induction of microspore embryogenesis in the Brassicaceae

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Abstract

The objective of this work was to enhance the quality and quantity of microspore-derived embryos of cruciferous species by using polyethylene glycol (PEG) to replace sucrose in the culture medium. The main advantage in using PEG is that it produces embryos that are morphologically more similar to zygotic embryos and have enhanced germination capabilities. When microspores were cultured in full strength NLN medium supplemented with 25% (w/v) PEG, the addition of 3 ml of full strength NLN with 13% (w/v) sucrose at 14 d was beneficial for embryo quality and quantity. Experiments showed that this PEG system could be used for a number of Brassica napus cultivars, as well as a number of other cruciferous species. PEG enhanced microspore embryogenesis in B. nigra, Crambe abyssinica, and Raphanus oleifera. Microspore-derived embryos were obtained from all cruciferous species evaluated (B. alboglabra, B. carinata, B. juncea, B. rapa, B. nigra, R. oleifera, Crambe abyssinica, Sinapis alba) using either sucrose or PEG as the osmoticum. Microspore embryogenesis was induced in B. napus in PEG-based cultures without a 32°C heat shock (i.e., 4, 15, 18, and 24°C). These temperature conditions were non-inductive when sucrose was used as the osmoticum. Spontaneous chromosome doubling occurred in 64–92% of the regenerated plants when PEG was used in the NLN culture medium, whereas in culture medium containing sucrose, the spontaneous doubling rate was 2–18%.

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Acknowledgment

The authors acknowledge the Saskatchewan Agriculture and Development Fund for their financial support. Technical support was provided by D. Stevenson and N. Di Santo.

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  1. Plant Biotechnology Institute, 110 Gymnasium Place, Saskatoon, Saskatchewan, S7N 0W9, Canada

    A. M. R. Ferrie & W. A. Keller

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  1. A. M. R. Ferrie
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Correspondence to A. M. R. Ferrie.

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Editor: K. D’Halluin

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Ferrie, A.M.R., Keller, W.A. Optimization of methods for using polyethylene glycol as a non-permeating osmoticum for the induction of microspore embryogenesis in the Brassicaceae . In Vitro Cell.Dev.Biol.-Plant 43, 348–355 (2007). https://doi.org/10.1007/s11627-007-9053-6

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  • DOI: https://doi.org/10.1007/s11627-007-9053-6

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