木立芦荟（Aloe arborescens Mill.）离体快速繁殖技术的研究

【作者】 傅术琳；

【导师】 程智慧； 周新民；

【作者基本信息】 西北农林科技大学 ， 蔬菜学， 2001， 硕士

【摘要】 芦荟是百合科芦荟属常绿草本植物，具有重要的医疗、保健美容价值。但因为授粉不亲和，不能用种子繁殖且常规繁殖率低，因此对芦荟采用组织培养的方法进行快速繁殖具有重要的意义。本研究以木立芦荟（Aloe arborescens Mill）为材料，研究了组织培养的全过程，探讨了影响快繁的一些因素，并提出了克服方法，初步建立了一套较完整的芦荟快繁体系。主要结果如下： 在茎尖、茎段、叶片和根这四种外植体中，不定芽和愈伤组织的分化能力存在明显差异。茎尖和茎段能直接诱导分化不定芽，叶片和根则不能形成不定芽。其中茎尖直接诱导不定芽的能力最高，每个茎尖能分化出不定芽19～20个，茎段只能分化8～9个不定芽。茎尖、茎段和幼嫩叶片基部均能诱导产生愈伤组织，而根则不能。 培养基不同，培养物的分化和发育也存在明显差异。在所选的MS，B5，改良MS（MS大量元素+B5微量元素和有机物），化肥培养基，马铃薯培养基等五种培养基中，以改良MS培养基对芽的分化发育最好，不定芽增殖倍数达12，显著高于其它培养基，其次是MS培养基，改良的化肥培养基也能获得良好的增殖效果，并且能够节约成本，在产业化快繁上可考虑采用。B5，马铃薯培养基和改良前的化肥培养基这三种培养基使组培苗生长发育受到抑制，并出现褐化和玻璃化现象。另外培养基中蔗糖浓度低于1.5％，易出现玻璃化现象，3％～4.5％浓度的蔗糖不但不定芽的分化能力高，而且对预防玻璃化现象也有一定的作用，琼脂浓度以0.7％为最好。 细胞分裂素6-BA对芽分化效果，以6-BA为最好，其适宜浓度随外植体的不同而有差异。茎尖和不定芽的增殖要求3.0mg.l^-1的6-BA，而茎段则为4.0mg.l^-1，但高浓度的6-BA对芽的伸长有抑制作用。KT对不定芽的分化效果不明显，但对芽的生长有促进作用。 低浓度的生长素NAA0.2mg.l^-1或IAA0.3mg.l^-1有利于不定芽的分化，而高浓度的生长素NAA2.0mg.l^-1或3.0mg.l^-1则有利于愈伤组织的诱导，但形成的愈伤组织却不能出芽。而只有在低浓度的6-BA和低浓度的生长素下诱导形成的愈伤组织才能产生不定芽，即在MS+6-BA0.5+2,4-D0.5+NAA0.4+ABA0.5+CH400+0.2％G中产生的愈伤组织具有不定芽分化的能力，愈伤组织和不定芽的诱导率分别为15％和5％。 生长调节剂B9能极大的促进不定芽的增殖，100mg.l^-1B9使不定芽的增殖倍数达11.5。500mg.l^-1CC明显促进不定芽的生长。3.0mg.l^-1PP₃₃₃使芽停止生长并白色愈伤组织化，但这种现象可被较高浓度的细胞分裂素解除并能继续生长和分化。 添加88mg.l^-1～132mg.l^-1的CaCl₂和用微孔滤膜封口可显著减轻玻璃化现象的发生。15mg.l^-1 硝酸银可减轻褐变的发生。 在生根方面，2．0＊g．！”1＊A和1刀＊g．卜＊＊A均明显地促进生根，根数可达个5条。1o＊s 和 1／3MS的低浓度的无机盐也有利于根的诱导。光照对生根不利，培养基中添加 0．2％的活性炭 而不加生长素对促进生根的效果明显，平均每个不定芽生根数为2．4条。 将生根的组培苗移栽到由2有机厩肥：l园田土或2＠石：l园田土组成的基质中并用透气 性好的覆盖材料微孔滤膜封口，不但极大的提高组培苗的移栽成活率使达到95％，而且使驯化和 移栽缩为一步，从而简化了快繁程序。更多还原

【Abstract】 In this study, the whole process of micropropagation of Aloe arborescens Mill in vitro was conducted .The main results shows as follows: There lay differences in differention among different explants . Adventitious buds were induced directly from both meristem and stem segments ,but not induced from leaves and roots . About I 9?0 and 8? buds were obtained from each meristem and stem segments , respectively. Calli formed from meristems , stem segments and leaf base ,but not from roots. MS , B5 , modified MS (MS macro salts and B5 micro salts and vitamins) ,fertilizer for nutrient solution and potato juice instead of MS inoganic salts were used . Results showed that modified MS had the best effects on differention of adventitious buds , each shoot achieving 12 buds ; MS were the secondly best medium ; 2105mg.l?fertilizer plus 44 mg.r?CaCI2 also facilitated bud multiplication , so this is of commercial importance in micro propagetion of Aloe in vitro. B5 and potato juice instead of MS inoganic salts were not suitable to growth and differentiation of shoots. The concentration of sucrose less than 1.5% resulted more serious vitrification , only 3.O0/o挆4.S% was optimal to differentiation and growth of adventitious buds . Appropriate Agar concentration was 0.7%. Different cytokinins exhibited different effects on differentitiation of cultures . High concentration of 6-BA induced the formation of adventitious buds greatly, but inhibited shoots elongation . 3.Omg.r 6-BA was most suitable to meristem differentitiation and multiplication of adventitious buds , but induction of buds from stem segments needed 4.Omg.U?-BA . Kinetin had no obvious effects on the induction of buds , but promoted bud growth . Low concentration of auxins were necessary to the induction of adventitious buds , 0.2mg.r NAA and 0.3 mg.UAA were optimal , respectively. Calli induced from explants cultred in media supplemented with 2.0 mg.[扤AA or 3.0 mg.r?,4-O alone couldn’t differentiated shoots, only those calli formed in MS + 2,4-D0.5 + 6.-BAO.S + NAAO.4 + ABAO.5 + CH400 + 0.2%G were induced adventitious buds , but only 5% of thse calli differentiated buds. Plant growth regulators I OOmg.V?B9 promoted multiplication of shoots greatly and 11.5 buds were obtained from each shoot . Shoots growed faster in medium containing 500 mg.l?CC (choline chloride) . 3.0 mg.1 PP333 stopped shoots growing and made them turn to white calli , but when subcultured in media with high concentration of cytokinins , shoots could continue to grow and differentiate. Addition of 88 mgfI 32 mg.raCI2 to media and sealing bottles with micropore embranes allieviated vitrification significantly . lSnigY?AgNO3 inhibited brown effectively , but at the same time made shoots grow slowly. Both 1.0 mg.[AA and 2.Omg.UBA induced 4--S pieces of roots . Low concentration of MS inorganic salts such as 1/2 MS and l/3MS had better effects on root formation than higher concentration of MS inorganic salts . Addition of activated charcoal to media promoted the differentitiation of root even without auxins in media. Shoots cultured in media solidified with sands growed vigiously and rooted , So sands could replace agar as supporter of shoots . This can reduce the cost of shoots multiplicated in vitro. Transplanting rooted shoots to medium composed with 2 manure : I soil or 2 vermiculite : I soil and covering pots with micropore embranes not only greatly increased survival rate of shoots to 95%, but更多还原