摘要
兰科植物独蒜兰Pleione bulbocodioides(Franch)Rolfe是生药山慈姑的基原植物，为常用中药，其假鳞茎具有清热化痰、解毒、消痈散结的作用，现已临枯竭而成濒危物种。为避免山慈姑(独蒜兰)资源灭绝，该论文研究解决山慈姑(独蒜兰)的人工繁殖技术，为山慈姑(独蒜兰)的可持续利用奠定基础。本论文以山慈姑(独蒜兰)球茎和种子为外植体，进行组织培养和植株再生系统的研究结果如下：
(一)以山慈姑(独蒜兰)球茎为外植体，比较附加不同浓度的6-BA、2，4-D和NAA的MS培养基对原球茎的诱导、芽的形成以及原球茎生根的影响。结果表明：以“MS+2 mg l~(-1)2，4-D+0.5～1.0 mg mg l~(-1)6-BA+0.05％干酪素+0.5％活性炭+3％蔗糖+0.55％琼脂”为培养基，能够很好地诱导外植体脱分化，并促进脱分化后的外植体分化形成新的原球茎，再生原球茎在1／2MS+0.2 mg l~(-1)NAA+0.5％活性炭+2～3％蔗糖+0.55％琼脂的培养基上诱导生根，然后发育形成根系发达、茎干粗壮的幼苗。
(二)以山慈姑(独蒜兰)的成熟种子为材料，比较不同的前期处理、不同的培养基及培养条件诱导种子萌发、萌发种子的分化成苗及试管苗移栽的影响。结果表明：山慈姑(独蒜兰)丛生芽和原球茎的诱导及分化与材料处理、培养基状态和激素组合有关。在(1)“MS+2.0 mg l~(-1)6-BA+0.2 mg l~(-1)NAA+3％蔗糖+0.50％琼脂，pH5.2～5.6”；(2)“MS+2.0 mg l~(-1)2，4-D+0.5 mg l~(-1)6-BA+3％蔗糖+0.50％琼脂+1.0％活性炭+0.05％干酪素，pH5.2～5.6”培养基中种子萌发率都高达40％，在(1)培养基中，种子先萌发形成小球体，再由小球体发育成幼苗；在(2)培养基中种子先形成原球茎，部分原球茎直接发育成幼苗，部分原球茎通过丛生芽途径再发育成小苗。在(2)号培养基中发育成的小苗比在(1)号中的健壮，茎粗；在(3)“1／2MS+0.2 mg l~(-1)NAA+2-3％蔗糖+0.2-1％活性炭+0.45-0.50％琼脂”培养基则有利于独蒜兰再生植株小苗生根进一步生长发育。
Pleione bulbocodioides (Franch) Rolfe is one of the most famous orchids distributed in a few countries in southeast and south Asia. It is mainly used as a decorative plant and in Chinese traditional medicine. However, its numbers are steadily declining, due to a lower rate of propagation in nature and over exploitation. So, it is essential that measures are taken to conserve and propagate this endangered orchid species. Therefore, an in vitro propagation technique could be a useful approach for the mass scale propagation of this orchid for commercial purpose. Two regenerated systems were established using bulbs and seeds as explants.
The effects of 6-BA, 2,4-D and NAA on protocorm induction, shoot formation and rooting of plantlets were evaluated using bulbs of P. bulbocodioides as explants. It can be seen from our results. Explants dedifferentiated and subsequently differentiated to form protocorms when cultured on MS medium supplemented with 2 mg 1~(-1) 2,4-D, 0.5～1.0 mg 1~(-1) 6-BA , 0.05% casein, 3% sucrose and 0.5% active carbon. These regenerated protocorms developed into well rooted plantlets as transplanted onto 1/2MS medium having 0.2 mg 1~(-1) NAA.
The effects of the different time of pretreatment and plant growth regulators on seed germination, formation and transplantation of plantlets were evaluated when seeds of P.bulbocodioides used as explants. Our result shows that the morphogenesis of explants is related to the original explants, basal medium and plant growth regulators. There were two pathways of plant regeneration when cultured on MS medium with different plant growth regulators. Seeds germinated into globular granules and continued to develop into plantlets when cultured on MS medium with 2 mg 1~(-1) 6-BA, 0.2 mg 1~(-1) NAA and 3% sucrose. Seeds germinated into protocorms when cultured on MS medium with 2 mg 1~(-1) 2,4-D, 0.5 mg 1~(-1) 6-BA, 3% sucrose and 0.05% carbon. Some of these protocorms directly developed into intact plantlets and others developed into plantlets through shoot formation on the same medium. Though the frequency of seed germination were over 40% when cultured on these two medium, physiological conditions of regenerated plants were different. The plantlets were stronger when cultured on MS medium with 2 mg 1~(-1) 2,4-D, 0.5 mg 1~(-1) 6-BA, 3% sucrose, 1% active carbon than cultured on MS medium with 2 mg 1~(-1) 6-BA, 0.2 mg 1~(-1) NAA and 3% sucrose. Well developed plantlets were obtained when transplanted into 1/2MS medium with 0.2 mg 1~(-1) NAA, 2～3% sucrose, 0.2～1.0% active carbon.
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