In this experiment, a Cystain UV precise P kit manufactured by Partec, Münster, Germany is used in extraction and staining of leaf cell nuclei, and the procedure of extraction and staining is processed on an ice bath filled with ice fragments. Firstly, 30-40 mg of newly developed leaves of the regenerated plants previously obtained in the experiment in accordance with the present invention (as best shown in FIGS. 1A-1C and 2, and as disclosed in the section “A” above) are cut off and placed in a flat-bottomed glass Petri dish (having a diameter of 5.5 cm), and 100 μl of extraction buffer is added in the Petri dish. Subsequently, the newly developed leaves are cut into fragments less than 1 mm by a double-sided razor blade, and are mixed with 400 μl of DAPI staining buffer in the Petri dish. The leaf fragment mixtures containing leaf cell nuclei are filtered through a filter having a mesh size of 30 μm. Accordingly, the filtrates are used for chromosome ploidy analysis in the subsequent procedure.
In this experiment, a Cystain UV precise P kit manufactured by Partec, Münster, Germany is used in extraction and staining of leaf cell nuclei, and the procedure of extraction and staining is processed on an ice bath filled with ice fragments. Firstly, 30-40 mg of newly developed leaves of the regenerated plants previously obtained in the experiment in accordance with the present invention (as best shown in FIGS. 1A-1C and 2, and as disclosed in the section “A” above) are cut off and placed in a flat-bottomed glass Petri dish (having a diameter of 5.5 cm), and 100 μl of extraction buffer is added in the Petri dish. Subsequently, the newly developed leaves are cut into fragments less than 1 mm by a double-sided razor blade, and are mixed with 400 μl of DAPI staining buffer in the Petri dish. The leaf fragment mixtures containing leaf cell nuclei are filtered through a filter having a mesh size of 30 μm. Accordingly, the filtrates are used for chromosome ploidy analysis in the subsequent procedure.
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