| Anthurium andraeaum is one of the perennial herb flowers which belong to Anthurium genus of Araceae family. It is widely used as cutting flower material and potted ornamental plant because of its excellence like various colors, peculiar flower shape, long florescence. It has a very high ornamental and commercial value. Recently, with the rapid development of Anthurium andraeaum industry, the dramatical needs for diverse Anthurium andraeaum varieties is becoming more and more obvious. In order to obtaining new Anthurium andraeaum germplasm and enrich breeding materials, colchicine and Ethyl methane sulfonate(EMS)were used as mutagenic agents, Anthurium andraeaum callus as test material in this paper to study the influence on the growth and differentiation of callus and the mutagenic effect of the two mutagenic agents. The mutation materials were identified and analyzed, the main results of the study were as follows.1. Polyploid induction of Anthurium andraeaum in vitro. The calli of Anthurium andraeanum were treated with colchicine at concentrations of 0.2, 0.3, 0.45 g/L for 3, 6, 9, 14 d respectively to induce polyploid plants. The optimal treatment was 0.3 g/L colchicine for 9 d with mutation rate of 60 %. There were 9 kinds of regenerated plantlets that were different from those of control in morphology. Polyploid plants grew slower and had thicker and deeper green leaves, shorter and thicker stem.2. Ploidy identification of Anthurium andraeaum polyploid. Root tip squash method obserbation showed that there were tetraploid cells in mutants and the chromosome number of tetraploid cells was 2n=4x=60, while that of the control was 2n=2x=30. Identification of flow cytometry showed that WPB buffer and Tris·MgCl2 buffer were the appropriate nuclear isolation buffers having more nuclei and very low CVs, tetraploids identified by root tip squash method resulted in histogram with two prominent DNA peaks of PI fluorescence at channels 180 and 360, while diploids resulted in histogram with a single peak of PI fluorescence at channel 180. The flow cytometry analysis results showed that these regenerated plantlets with tetraploid cells were mixoploids.3. EMS mutagenesis of Anthurium andraeaum in vitro. The calli of Anthurium andraeanum were treated in vitro with EMS at concentrations of 0.2, 0.4, 0.6 % for 12, 24, 36 h respectively to induce genetic mutation plants. It was found that EMS treatment had great effect on the growth of callus. Some calli got brown and died, which was related to the intensity of EMS treatment. Under conditions of this experiment, the better EMS treatment for Anthurium andraeanum callus was 0.6 % EMS treated for 12 h and its death rate was close to the half lethal dose. There were many regenerated plantlets treated with EMS that were different from those of control in morphology and 13 mutants were acquired.4. RAPD analyzing of Anthurium andraeaum regenerated plantlets treated with EMS. 48 regenerated plantlets treated with EMS and the control were amplified by PCR method. The results indicated that 12 arbitrary primers amplified 55 bands ranging from 100 to 2000 bp with an average of 4.6 bands per primer, among them 21 bands were polymorphic and the percentage was 38 %. Compared the bands of regenerated plantlets with those of control, there were differences among them and DNA polymorphisms were observed. The results indicated that the regenerated plantlets had genetic changes. |
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