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From cells to embryos to rooted plantlets in a mist bioreactor

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Abstract

A mist bioreactor using a disposable bag as culture chamber was used to propagate carrot embryogenic cells into rooted plantlets. The best operating configuration was akin to a vertical hanging garden using 50–90 μm nylon mesh for explant attachment. Cells spray inoculated into the reactor were 51.2 % viable. Misting cycle and aeration conditions were studied and showed that under the same hourly volumetric nutrient feed and 0 VVM, embryo development in the reactor was best using a 0.3 min on/2.7 min off misting cycle, yielding about 23 % post heart stage embryos. Compared to 0 VVM, 3 % CO2 enrichment improved embryo development in reactor culture. Spray inoculated cells also attached to several vertically hung poly-l-lysine coated strips and then developed in situ into embryos. Cell attachment was significantly improved when they were suspended in salt-free sucrose solution during spray inoculation. Almost 90 % of the originally attached cells remained on the nylon mesh 24 h later after spraying with B5 medium in the mist reactor. Strip grown embryos had the same post heart stage ratio but shorter overall length compared to those developed on a horizontal platform. Young plantlets developed uniformly up and down the hanging strips and did not detach after 3 weeks of culture suggesting this technology may prove useful for improving micropropagation.

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Abbreviations

N50, N70, N90:

Nylon screen/mesh with openings of 50, 70 and 90 μm, respectively

P74, P105:

Polypropylene screen/mesh with opening of 74 and 105 μm, respectively

PLL:

Poly-l-lysine

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Acknowledgments

The authors thank WPI for supporting L Fei. Advice from Dr. Melissa Towler and Prof. Elizabeth Ryder of the Department of Biology and Biotechnology at WPI was greatly appreciated.

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  1. Department Biology and Biotechnology, Worcester Polytechnic Institute, 100 Institute Rd, Worcester, MA, 01609, USA

    Liwen Fei & Pamela J. Weathers

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  1. Liwen Fei
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  2. Pamela J. Weathers
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Correspondence to Pamela J. Weathers.

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Fei, L., Weathers, P.J. From cells to embryos to rooted plantlets in a mist bioreactor. Plant Cell Tiss Organ Cult 116, 37–46 (2014). https://doi.org/10.1007/s11240-013-0380-5

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