抗性湿地松体胚的发育、成熟与萌发

为加快抗松针褐斑病湿地松体细胞胚胎发生技术的产业化应用进程,本研究以肌醇浓度、脱落酸(ABA)的添加方式、基本培养基、ABA与聚乙二醇(PEG6000)互作、PEG6000及琼脂糖种类及浓度、活性炭(AC)、ABA、液体悬浮培养等因素对抗性湿地松体胚的发育、成熟与萌发的影响进行研究。结果表明:添加8g/L的肌醇为宜,过高浓度的肌醇则不利于湿地松体胚的发育；ABA可以采用高压灭菌；基本培养基、ABA、PEG6000互作对湿地松体胚的成熟影响很大,基本培养基以LP为最佳,ABA的最适浓度为10mg/L,PEG6000以添加25g/L、50g/L为最佳；在湿地松体胚的成熟培养过程中,PEG6000的浓度不能高于100 g/L；在湿地松体细胞胚的成熟培养基中,添加60g/L蔗糖最合适；湿地松体细胞胚的成熟培养基中添加0.5g/L或1g/L的活性炭最有利于体胚的成熟；培养基的状态对体胚的高频率诱导具有一定的影响,悬浮培养以100mL的三角瓶装30mL的培养液,摇床转速以130r/min为宜,液体转固体时吸取1mL培养液为宜；未建立成熟的体胚发生体系前,以采用固体培养为佳。最佳成熟培养基、激素及部分添加物组合为LP+10mg/L ABA+50g/L PEG6000+60g/L蔗糖+1g/L活性炭。

Development, Maturation and Germination of Resistant Slash Pine Somatic Embryos

In order to speed up the process of industrialization and application of Slash Pine somatic embryogenesis technology against pine needle brown spot, the influence of somatic embryo development, maturation and germination of resistant slash pine such as the concentration of inositol, the method of adding abscisic acid (ABA), the basic medium, ABA, and polyethylene glycol (PEG6000), activated carbon (AC), liquid suspension culture were studied. The results showed that it was advisable to add 8g/L inositol, and too high concentration of inositol was not conducive to the development of slash pine somatic embryos; ABA could be autoclaved; basic medium, ABA, PEG6000 interacted with slash pine somatic embryos maturation had a great influence. LP was the best basic medium, and the optimum concentration of ABA was 10mg/L. For PEG6000, 25g/L and 50g/L were the best; during the maturation and culture of slash pine somatic embryos, the concentration of PEG6000 should not be higher than 100 g/L; adding 60g/L sucrose was the most suitable in the maturation medium of slash pine somatic embryos ; Adding 0.5g/L or 1g/L of activated carbon to the maturation medium of Slash pine somatic embryos was most conducive to the maturation of somatic embryos; the state of the medium had a certain effect on the high-frequency induction of somatic embryos, for suspension culture, 30 mL of culture fluid was filled in a 100 mL Erlenmeyer flask, and the appropriate shaker speed was 130r/min,When liquid was changed to solid, 1 mL of culture medium was appropriated; it was better to use solid culture before the establishment of a mature somatic embryogenesis system. The best combination of maturation medium, hormones and additives was LP+10mg/L ABA+50g/L PEG6000+60g/L sucrose+1g/L activated carbon.