Isolation and Characterization of Microprotoplasts from Propyzamide-treated Cell Suspension Cultures of Hemerocallis hybrida.
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Accession number;02A0274320
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| Title;Isolation and Characterization of Microprotoplasts from Propyzamide-treated Cell Suspension Cultures of Hemerocallis hybrida. |
| Author;
SAITO H
(Niigata Univ., Niigata, Jpn)
NAKANO M
(Niigata Univ., Niigata, Jpn)
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Journal Title;Breed Sci
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Journal Code:Y0311B
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ISSN:1344-7610
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VOL.52;NO.1;PAGE.51-56(2002)
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| Figure&Table&Reference;FIG.3, TBL.1, REF.17 |
| Pub. Country;Japan |
| Language;English |
| Abstract;To produce intergeneric hybrid plants with one or a few chromosomes via microprotoplast fusion in the Liliaceous ornamentals, we developed a system for isolating microprotoplasts from micronucleated cells in suspension cultures of Hemerocallis hybrida cv. Stalla d'Oro(2n=2x=22). Micronucleation was efficiently induced with a micronucleus index up to 19.1% by the following sequential treatment of suspension cultures: initially with 2mM hydroxyurea for 24h and then with 8.MU.M propyzamide for 60h with the addition of 20.MU.M cytochalasin-B to the cultures 20h after the initiation of the propyzamide treatment. Following the enzyme treatment of the micronucleated cells and ultra-centrifugation with a continuous iso-osmotic gradient of Percoll solution, microprotoplasts were isolated, each of which had a small nucleus surrounded by a thin rim of cytoplasm. Microprotoplasts less than 10.MU.m in diameter were obtained with yield of 2.9*104 cells per 1ml packed cell volume of suspension cells through sequential filtration with nylon sieves with decreasing pore sizes(50, 20 and 10.MU.m). The DNA content of the microprotoplasts was less than the 2C level as indicated by flow cytometric analysis, and the relative fluorescence intensity in some of their nuclei corresponded to one or a few chromosomes. (author abst.) |
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