金钱松种子萌发条件优化和快速繁育研究

发布时间：2019-03-30 21:41

【摘要】：实验通过单因素试验和正交试验的研究方法,优化金钱松种子萌发条件;建立金钱松快速繁育体系;建立金钱松植物细胞的悬浮培养和植株快速再生的培养体系;探究利用金钱松种子诱导多倍体的最佳培养条件;从金钱松植株的新鲜枝条中分离出内生菌,并进行了抗菌试验研究。具体实验结果如下:(1)通过单因素试验研究了1%多菌灵溶液消毒处理对其种子的萌发率和染菌率的影响。实验结果表明,随着1%的多菌灵溶液处理金钱松种子时间的增长,其种子的萌发率和染菌率均表现出逐渐下降的趋势,最有利于提高金钱松种子的萌发率和降低其种子的染菌率的处理方式为1%多菌灵溶液处理金钱松种子3h,种子的萌发率为(82±0.32)%,而污染率仅为(3±0.49)%。另外,实验还研究了不同赤霉素浓度、不同温度和浸泡不同时间对金钱松种子萌发的影响,研究结果表明,金钱松种子在60℃下,用6‰赤霉素浸泡2 h后,再放在培养皿中培养30 d,种子萌发率最高,为97.33%。(2)以金钱松胚轴为外植体诱导愈伤组织的最佳培养基为MS+2,4-D 1.0 mg/L+6-BA 0.5 mg/L+KT 0.1 mg/L;影响胚轴外植体快速产生愈伤组织除了激素配比外,还与外植体的接种方式、切割方式有关;研究培养过程中形成的六种愈伤组织,得出颜色呈绿色、浅绿色的愈伤组织细胞活力是最高的。(3)金钱松细胞悬浮培养的最佳培养基MS+6-BA 1.0 mg/L+2,4-D 0.3 mg/L+蔗糖40 g/L+接种量1.2 g/40 ml+p H 6.0;在细胞的生长过程中,具体表现为,延迟期为0~3 d,指数期为3~7 d,衰落期为7~20 d;另外,在实验中观察还发现,翠绿的愈伤组织生长速度快,黄绿色和黄色的愈伤组织生长速度慢、且褐变速率快。(4)在金钱松植株快速再生研究中,选取金钱松叶片作为外植体诱导愈伤组织,其愈伤组织诱导的最佳培养基为GD+NAA 1.0 mg/L+GA31.0 mg/L,在培养30 d后统计愈伤组织诱导率为65.4%,诱导不定芽最佳培养基为MS+TDZ 0.2 mg/L+Ad 20 mg/L,培养35 d诱导率为23.1%;诱导不定根的最佳培养基为1/3MS+IAA 1.0 mg/L+活性炭30 mg/L,培养35 d诱导率为54.0%。无菌苗在室内环境下开盖培养三天以后栽种在大棚中,成活率为93%。(5)在开展金钱松植株的多倍体诱导过程中,最佳诱导培养条件为,在30℃下,0.2%的秋水仙素浸泡36 h,诱导率为78.63%;而诱导率最低的条件为,在20℃下,0.1%的秋水仙素中浸泡12 h,诱导率为49.94%;当秋水仙素浓度0.2%,秋水仙素作用时间从12 h升高到36 h时,金钱松多倍体诱导率逐渐上升,当作用时间从36 h升高为48 h时,诱导率有下降趋势。(6)本研究从金钱松新鲜枝条中分离出了JJ18、JJ243、JJ314、JJ318、JJ416、JJ27和JJ46共七内生菌;用金黄色葡萄球菌为指示菌,筛选出具有抗菌作用的内生菌;提取内生菌JJ18代谢物,作用于白色念珠菌、红色毛癣菌、酿酒酵母菌和黑曲霉,测定了各种菌的MIC(最小抑菌浓度)和MBC(最小杀菌浓度)。
[Abstract]:By means of single factor experiment and orthogonal experiment, the germination conditions, rapid propagation system, suspension culture system and plant regeneration culture system of Pinus pinnata were optimized, and the results were as follows: (1) the seed germination conditions of Pinus pines were optimized, and the results were as follows: 1. To explore the optimum culture conditions for inducing polyploidy by using the seeds of Pinus mongolica, endophytic bacteria were isolated from fresh branches of Pinus koraiensis, and the antibacterial test was carried out. The results were as follows: (1) the effect of disinfection treatment with 1% carbendazim solution on the germination rate and infection rate of the seeds was studied by single factor experiment. The results showed that with the increase of 1% carbendazim solution treatment time, the germination rate and the infection rate of the seeds decreased gradually, and the seed germination rate and the infection rate decreased gradually with the increase of the time of 1% carbendazim solution treatment. The best treatment was 1% carbendazim solution for 3 h, the germination rate was (82 卤0.32)%, and the contamination rate was (3 卤0.49)%. In addition, the effects of different concentration of gibberellic acid, different temperature and different time on seed germination were studied. The results showed that the seeds were soaked in 6 鈥，

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