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从印度娃儿藤节间外植体获取愈伤组织,分析了糖、赤霉素(GA3)及脱落酸(ABA)对愈伤组织形成体细胞的影响。实验证明,含4μmol/L2,4-二氯苯氧乙酸(2,4-D)的MS培养基是获得具有成胚功能的愈伤组织的最佳培养基。在含有6μmol/L激动素(Kn)的MS培养基上,高达69%的愈伤组织分化为体细胞胚,平均单位外植体(每克愈伤组织)产胚25个。在6μmol/LKn存在的条件下,分析了蔗糖、葡糖糖对胚产生的影响,不同的糖及不同糖浓度对体细胞胚的发生影响很大。6μmol/L Kn与200mmol/L蔗糖处理胚胎发生率最大(71%),单位外植体生成49个胚。然而葡萄糖与Kn、或者葡糖糖、蔗糖与Kn三者加在一起则降低成胚率及产胚数。一定浓度的GA3和ABA能促进体细胞胚的产生。在含200mmol/L蔗糖的培养基中加10μmol/LGA3胚的生成率为98%,单位外植体产胚51个。在含200mmol/L蔗糖的培养基中加2μmol/L ABA能显著增加体细胞胚的量,该培养基上每外植体平均生成44个胚,产率为95%。本研究显示,含200mmol/L蔗糖的培养基中分别加入6μmol/L Kn、10μmol/L GA3或者2μmol/L ABA能显著提高印度娃儿藤体细胞胚发生率,而单独的葡萄糖或葡糖糖和蔗糖则有抑制作用。得到的胚均能正常发育并分化为植株。
The callus was obtained from the explants of V. villosum and the effects of sugar, gibberellin (GA3) and abscisic acid (ABA) on the callus formation of somatic cells were analyzed. Experiments show that MS medium containing 4μmol / L 2,4-dichlorophenoxyacetic acid (2,4-D) is the best medium for obtaining embryogenic callus. On MS medium containing 6 μmol / L kinetin (Kn), up to 69% of callus differentiated into somatic embryos with an average of 25 explants per gram of explant (per gram of callus). In the presence of 6μmol / LKn, the effects of sucrose and glucose on embryos were analyzed. Different sugars and different sugar concentrations had a great effect on somatic embryogenesis. The embryogenesis rate was the highest (71%) with 6μmol / L Kn and 200mmol / L sucrose, and 49 embryos per explant. However, the combination of glucose and Kn, or glucose, sucrose and Kn reduced the embryo formation rate and embryo production. A certain concentration of GA3 and ABA can promote the production of somatic embryos. In the medium containing 200mmol / L sucrose, the rate of 10μmol / L GA3 embryo formation was 98%, and the number of explants producing embryos was 51. The addition of 2μmol / L ABA to medium containing 200mmol / L sucrose markedly increased the amount of somatic embryos, with an average of 44 embryos per explant in this medium with a yield of 95%. The results showed that the addition of 6μmol / L Kn, 10μmol / L GA3 or 2μmol / L ABA to medium containing 200mmol / L sucrose significantly increased the embryogenic rate of somatic embryo in India, while glucose or glucose alone And sucrose is inhibited. The embryos obtained can all normally develop and differentiate into plants.