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Isolation of Protoplasts from Tissues of 14-day-old Seedlings of Arabidopsis thaliana
Published on: August 17, 2009
[Isolation and properties of the protoplasts of Streptomyces erythraeus]
Abstract:
The paper is concerned with development of conditions for cultivation of the erythromycin-producing organism and preparation and maintenance of its stable viable protoplasts. Optimal conditions for the culture growth and protoplasting were developed. Two-stage cultivation of the organism on media PB and S provided dense diffuse or diffuse local growth characterized by low differentiation and higher homogenicity. The incubation time at stage I was 66 hours and that at stage II was not more than 24 hours at respective temperatures. The culture was incubated on a shaker in the presence of glycine at the minimum concentration. The presence of glycine in the medium altered the culture cell walls which was evident from changing of staining by Gram from + to +/- . Treatment of such a culture with lysozyme for 30 minutes provided formation of up to 2.10(9) protoplasts per 1. ml. It is possible to maintain the protoplasts in frozen state at -20 degrees C in medium P for 1 month. Under such storage conditions the titer of the viable protoplasts as compared to the initial one decreased only 2-fold after the one-month storage.
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