Journal
PLANT BIOTECHNOLOGY
Volume 27, Issue 4, Pages 353-U73Publisher
JAPANESE SOC PLANT CELL & MOLECULAR BIOLOGY
DOI: 10.5511/plantbiotechnology.27.353
Keywords
Bamboo protoplasts; Bambusa vulgaris; Lithachne pauciflora; Phyllostachys meyeri; Sasa jotanii
Funding
- Ministry of Education, Culture, Sports, Science and Technology. [21380089]
- Collaboration Center for Research & Development Intellectual Property Center
- Utsunomiya University
- [20.7324]
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Protoplasts with a yield greater than 2.0 x 10(6) cells per 0.3 g fresh weight were isolated from leaves of four bamboo species: an herbaceous bamboo, Lithachne pauciflora, and three woody bamboos, Phyllostachys meyeri, Sasa jotanii, and Bambusa vulgaris. For aseptic protoplast isolation, folded leaves were sterilized in 70% ethanol, and the internal tissues were drawn out from the leaf sheath. The leaf material was incubated for 4 h in a modified White's medium solution containing 0.8 M mannitol ( pH 5.8, 32 degrees C) and an enzyme mixture containing 3% Macerozyme R-10, 2.5% Meicelase, and 2% Cellulase Onozuka for woody bamboos, and 2.5%, 2.5%, and 1% of these enzymes, respectively, for L. pauciflora. Protoplasts were filtered through a double-layered Miracloth funnel and then washed with 0.6 M to 0.4 M mannitol-containing White's medium ( pH 5.8) prior to analysis. Protoplast viability averaged 83% as determined by FDA staining.
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