马尿泡与山莨菪组织培养及马尿泡愈伤组织对低温胁迫的生理响应

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	马尿泡与山莨菪组织培养及马尿泡愈伤组织对低温胁迫的生理响应
其他题名	Tissue culture of Przewalskia tangutica & Anisodus tanguticus and physiological response of the Przewalskia tangutica embryognic calluses uhder low temperature stress
	徐文华
学位类型	硕士
导师	陈桂琛
	2005-06-08
学位授予单位	中国科学院西北高原生物研究所
学位授予地点	西北高原生物研究所
关键词	马尿泡山莨菪愈伤组织植株再生低温胁迫氧化损伤渗透调节抗氧化酶
摘要	本文以青藏高原药用植物山莨菪（Anisodus tanguticus （Maxim.）Pascher）和青藏高原特有属植物马尿泡（Przewalskia tangutica Maxim.）的休眠芽为材料，利用组织培养技术研究了两种药用植物通过器官培养分化芽的途径进行快速繁殖，分别建立了愈伤组织培养和植株再生体系，从而筛选出了有利于马尿泡和山莨菪芽分化、增殖培养、生根诱导以及愈伤组织细胞生长的系列最佳培养基，对马尿泡和山莨菪优质种苗繁育具有一定的潜在应用价值，并对濒危植物马尿泡种质资源的保存具有重要的生态学意义。以马尿泡胚性愈伤组织为实验材料，研究了我国青藏高原特有属特有种植物马尿泡对极端低温胁迫环境的不同生理生化响应，揭示了在低温胁迫下马尿泡细胞内活性氧损伤与渗透调节物质以及抗氧化酶之间的相互关系，探讨了离体培养细胞条件下马尿泡抗逆能力的保持性。主要获得以下结果：（1）采用马尿泡野生植株休眠芽为离体培养的初始材料，成功的建立了马尿泡愈伤组织诱导、植株再生及离体快速繁殖体系。主芽周围的侧芽为最佳外植体。芽分化最佳培养基为MS + 2.0mg/L6-BA + 1.0mg/LNAA，芽的分化率达100%，平均每个芽每代可增殖30个芽以上；生根诱导培养基为1/2MS附加0.5mg/LIBA，生根率达96%；愈伤组织诱导培养基为MS + 0.5mg/L6-BA + 0.5mg/LNAA + 1.0mg/L2,4-D,出愈率达100%。（2）山莨菪大田栽培植株上的休眠芽可做为离体培养的良好外植体，成功的建立了山莨菪愈伤组织诱导、植株再生及离体快速繁殖体系。芽分化最佳培养基为MS + 2.0mg/L6-BA + 1.0mg/LNAA + 3.0mg/LZT，芽的分化率达100%，平均芽数在10~15个；芽稳定增殖培养基为MS + 3.0mg/L6-BA+ 1.0mg/LNAA；生根诱导培养基为1/2MS+ 1.0 mg/LNAA +3%蔗糖，35d后的生根率达100%；愈伤组织诱导培养基为MS + 0.2mg/L6-BA + 0.2mg/LNAA + 2.0mg/L2,4-D，出愈率为93.2%，且愈伤组织启动期为最早，为接种后的第17d。（3）马尿泡胚性愈伤组织可做为低温胁迫下生理生化响应研究的良好供试材料，研究了离体细胞培养条件下马尿泡抗逆能力的保持性。低温胁迫下马尿泡愈伤组织不同程度的表现出被伤害和生长抑制，细胞内活性氧损伤与渗透调节物质以及抗氧化酶等相互之间的动态平衡可抵抗外界低温环境的胁迫。低温胁迫研究结果表明： ①马尿泡愈伤组织对零上低温的胁迫表现出一定的耐寒性，而零下低温会对其造成较严重的伤害。 ②低温胁迫下，马尿泡愈伤组织的相对含水量基本保持稳定，相对细胞活力随温度的降低显著下降。 ③低温胁迫下，马尿泡愈伤组织的MP、MDA含量、AOS含量都随温度的降低而显著升高。 ④渗透调节物质甘氨酸甜菜碱（glycine betaine）和游离脯氨酸（proline）的含量随温度的降低逐步升高，做出了积极的响应 ⑤马尿泡愈伤组织SOD、POD、CAT和APX四种酶的活性表现出一致的趋势，即先升高后降低。在活性氧对细胞造成过氧化损伤时都表现出较高的酶活性。
其他摘要	The calluses culture and plantlet regeneration system of the two kind of medicinal and endemic genera plants of Przewalskia tangutica Maxim. and Anisodus tanguticus （Maxim.）Pascher, distributed at the tibetan plateau, were established separately using the plant tissue culture technology. The dormant buds as experiment material , have carried on the rapid reproducation through the organ redifferentiation bud's way. Thus we screened out a series of the best culture medium being favor of bud differentiation , the multiplication culture, root induction and calluses cell growth. It has certain latent application value to high quality seedling breeding of the przewalskia tangutica and anisodus tanguticus, and has extremely important ecology significance to conservated endangered the species germplasm resources of przewalskia tangutica. Meanwhile, we, using embryognic calluses, studied the przewalskia tangutica plant different physiological response under the extreme low temperature stress environment, and detected the antioxidant enzymatic activities and the contents of osmolytes solutes and the reactive oxygen species. So we have revealed the correlation of reactive oxygen species and osmotic adjustment solute content as well as the activity of antioxidant enzymes, and have discussed the anti-stress ability's retentivity of the przewalskia tangutica plant under the condition of in vitro culture cell. The main results are summarized as follows: (1) Our experiment has successfully established a system for calluses induction, and plantlet regeneration and in vitro rapid propagation of przewalskia tangutica using the wild plant dormant buds for the initial material. The lateral buds around the main bud are best of explants. The bud differentiation best medium is MS + 2.0mg/L6-BA +1.0mg/LNAA, the differentiation rate reaches 100%; The medium for root induction is 1/2MS added with 0.5mg/LIBA, the rate to reaches 96%; Calluses inducing medium is MS + 0.5mg/L6-BA + 0.5mg/LNAA + 1.0mg/L2,4-D, the rates reach 100%. (2) A system was successfully established for calluses induction, and plantlet regeneration and in vitro rapid propagation of anisodus tanguticus, using the field cultivation plant dormant buds for the better explant materials. The bud differentiation best medium is MS + 2.0mg/L6-BA + 1.0mg/LNAA + 3.0mg/LZT, the differentiation rate reaches 100%, and the bud steadly multiplication medium is MS + 3.0mg/L6-BA + 1.0mg/LNAA; The medium for root induction is 1/2MS + 1.0mg/L NAA + 3%sucrose, the rate to reaches 96%; Calluses inducing medium is MS + 0.2mg/L6-BA + 0.2mg/LNAA + 2.0mg/L2,4-D, the rates reach 93.2%. (3) The przewalskia tangutica embryognic calluses was good material to studied different physiological response under the low temperature stress environment, and we have studied under the exsomatize cell culture condition the przewalskia tangutica resistance ability retentivity. Under the low temperature stress, the przewalskia tangutica calluses have varying degree the performance to injure and the growth suppression, in the cell the active oxygen damage and the seepage adjustment material as well as the antioxidase and so on the between dynamical equilibrium may resist the low temperature stress. And the findings indicated that: ①The przewalskia tangutica has certain resistance to cold to the zero in low temperature, but the below zero low temperature can cause the serious damage to it. ②Under the low temperature stress, the przewalskia tangutica cell relative water content is basic stable, but the relative cell vigor reduces along with the temperature drops. ③The research discovered that przewalskia tangutica relative electric conductivity (MP), the MDA content, and the active oxygen species (AOS) content were all obviously increased long with the temperature reduces under the low temperature stress. ④The content of glycine betaine and the proline of osmolytes solutes were gradually increased along with the stress temperature reduced, and has made the positive response. ⑤The antioxidant enzymatic activities demonstrate that SOD, POD, CAT, APX activeness displays the consistent tendency, i.e from rises to fall along with the temperature drops. Four kind of enzyme activeness increased along with the processing temperature drops in the zero low temperature stress, but under the below zero low temperature enzyme activeness reduces along with the processing temperature drops, and all displays the high enzyme activity when the active oxygen damag.
页数	95
语种	中文
文献类型	学位论文
条目标识符	http://210.75.249.4/handle/363003/3138
专题	中国科学院西北高原生物研究所
推荐引用方式 GB/T 7714	徐文华. 马尿泡与山莨菪组织培养及马尿泡愈伤组织对低温胁迫的生理响应[D]. 西北高原生物研究所. 中国科学院西北高原生物研究所,2005.