兰属、兜兰属、石斛属植物叶片的扫描电镜观察

对兰科植物的兰属、兜兰属及石斛属１６个种折叶片及其横断面进行了扫描电镜的观察。兰属各种叶片上表皮细胞均为矩形,上表皮细胞表面具小乳突或不明显突起。石斛属及兜兰属的各个种上下表皮细胞均为多边形,但石斛属表皮细胞表面无坦无纹饰,而兜兰属花叶类上表皮细胞表面明显呈乳突状,绿叶类呈龟背状隆起。兰属及石斛属叶片叶肉组织没有栅栏组织及海绵组织的分化,而兜兰属的绿叶类叶肉不分化;花叶类叶肉有分化。     

三种兰科植物的保护遗传学研究初探

采用随机扩增多态 DNA(RAPD)分析研究了中国3种珍稀濒危兰科植物硬叶兜兰(Paphiopedilum micranthum Tang et Wang)、麻栗坡兜兰(P. malipoense S.C.Chen et Tsi)和独花兰(Changnienia amoena Chien)的遗传多样性与群体遗传结构.12个RAPD引物在2种兜兰中共扩增出131条带.对4个硬叶兜兰群体的检测表明其物种水平的多态条带百分率(PPB)为 71.6%,Nei 的基因多样度(h)为 0.217 1,Shannon多样性指数 (I) 为 0.330 1;4个群体的平均多样性水平为 PPB = 45.2%,h = 0.145 7,I = 0.220 4,低于远交兰花的平均水平.在总遗传变异中,群体间遗传变异占20.31%,略高于远交物种的平均水平.在物种水平上,麻栗坡兜兰的PPB为49.5%,h为0.117 4,I为0.176 4,均大大低于硬叶兜兰.对11个独花兰群体采用16个RAPD引物共扩增出119条带.物种水平PPB=76.5%,h=0.194 1,I=0.305 8;在群体水平上,上述3个指标的平均值则分别为37.2%、0.119 7和0.181 0,均低于远交兰花的平均水平.群体间的遗传变异占45.27%,遗传分化明显高于远交物种的平均水平.导致3个物种遗传多样性偏低而群体间遗传分化较高的主要原因在于人为的过度采挖和生境的片断化.研究结果为兰花保护策略和措施的制定提供了理论基础.     

小叶兜兰的果实生长和雌配子体发育

为了解濒危兰科植物小叶兜兰(Paphiopedilum barbigerum Tang et Wang)胚珠和雌配子体的发育过程,采用常规石蜡切片技术对其果实的生长动态进行了研究。结果表明,授粉后60~75 d的蒴果内种子数量迅速增加,到授粉后120 d时种子充满整个蒴果。授粉后40 d的胎座上分化形成多数由1层表皮细胞包被1列细胞的胚珠原基;授粉后60 d时位于胎座指状结构末端处紧靠表皮细胞下方的孢原细胞分化为大孢子母细胞。之后,大孢子母细胞经过减数分裂和有丝分裂最终形成成熟胚囊;授粉后135 d胚囊发育成熟,附着在胎座上的种子个体分化明显。小叶兜兰胚囊的发育类型为双孢子葱型,胚珠为倒生胚珠,薄珠心,单珠被,成熟胚囊为8核。这为小叶兜兰的生殖生物学及繁殖体系的建立提供理论依据。     

兜兰胚胎学的研究

云南兜兰花药壁有五层细胞,绒毡层细胞具双核,属分泌型。小孢子母细胞减数分裂为同时型。四分体中四个小孢子呈四面体或左右对称式排列。成熟花粉为二细胞型。单粒分散的花粉包裹于黄色粘性物质中。生殖细胞最初形成的壁为胼胝质的,待游离到营养细胞质中时,具一层很薄的 PAS 正反应的壁,直到花药开裂时这层壁仍存在。成熟花粉无特化的萌发孔,只具薄壁区。胚珠为薄珠心,具一层珠被,胚囊发育为葱型,成熟胚囊为6—8核,胚发育过程中,具2—4细胞胚柄。胚乳具二核。种子成熟时胚柄及胚乳核都消失。成熟种子只具单层细胞的种皮和一个未分化的球形胚。     

珍稀濒危植物硬叶兜兰的遗传多样性及遗传结构研究

由于人为采集、走私贩卖以及生境的破坏,分布于中国西南石灰岩地区的野生硬叶兜兰居群受到严重的干扰与威胁。为有效地保护这种珍稀野生植物,本研究采用ISSR和SRAP两种分子标记对15个硬叶兜兰野生居群进行遗传多样性及遗传结构的研究。结果表明,硬叶兜兰在物种水平上具有较高的遗传多样性（ISSR：PPB=91．66％,He=0．3839;SRAP：PPB=99．29％,Hc=0．2806）。硬叶兜兰居群间存在一定程度的遗传分化（ISSR：Gs1： 0．2577;SRAP：Gst=0．2383）,可能由于较低的基因流（ISSR：Nm=0．7201;SRAP：Nm=0．7991）所致。UPGMA聚类分析以及主成分分析均把15个居群分成2个主要分支。居群间的地理距离和海拔差距是引起居群遗传分化的自然因素。     

小叶兜兰的种子发育和无菌萌发

为建立小叶兜兰的繁育技术体系,本研究通过无菌播种的方法,辅以TTc生活力测定等方法,比较了小叶兜兰种子在授粉后不同发育时期和培养条件下的萌发率,对小叶兜兰种胚的发育过程进行了显微观察,探讨种胚发育程度与萌发的关系。结果表明,小叶兜兰种胚的发育阶段对萌发的影响最大,授粉后255d的种子萌发率最高（90．71％）,该阶段种子仍呈白色但微干燥,种胚刚发育至球形胚阶段,胚柄尚存。1／4MS和1／2MS为小叶兜兰适宜的基本培养基,添加100mg·L^-1的土豆汁对小叶兜兰的无菌萌发有良好的促进作用。     

Conservation ecology of endangered species Paphiopedilum armeniacum (Orchidaceae)

Paphiopedilum armeniacum is an endangered orchid species, endemic to China. During the period of April 2000 to October 2005, 66 observation sites were selected in Luoshapo of Nushan Mountains in Yunnan, China, to carry out the conservation ecological research on P. armeniacum. A total of 443 genets (1302 ramets in total) of P. armeniacum were sampled, their biological characteristics such as reproductive pattern, phenology, and life cycle were observed, and the ecological habits of the species such as the habitat and the structure of communities were studied. Experiments on ex-situ conservation were conducted, and the cloned ramets were replanted to their original habitat after ex-situ reproduction in Shenzhen, Guangdong, China. The relationships between P. armeniacum and climate, vegetation, other environmental factors in the original habitat, and the biological characteristics of asexual offsprings of P. armeniacum, which were replanted to the original habitat after ex-situ cultivation and reproduction, were investigated. The studies show that P. armeniacum in Luoshapo grows very well in secondary shrub boskets or in tussocks on limestone hills. It has both sexual and asexual reproduction. Asexual reproduction serves to complete the sexual reproduction and to extend the lifetime of genets, while it does not reduce sexual reproduction. There are two modes of asexual reproduction-by tillering or by producing rhizomes. The litter of shrub boskets or tussocks provides P. armeniacum with humus, and the rhizome reproduction of P. armeniacum is an adaptation to the litter-covered condition, i.e. to escape from the unfavorable environment. Blooming rate of ramets is 7.39% 1.02%, and fruit set rate from the blooming ramets is 32.23% 12.08%. P. armeniacum is able to invade the moderately destroyed forests and those in early restoring but is unable to grow in large dense forests. P. armeniacum also grows very well in artificial spare woods in Shenzhen and can reproduce many cloned ramets, which can normally bloom and yield fruits after being replanted to the original habitat. The results of this study show that P. armeniacum can be conserved by ex-situ conservation and by replanting the ex-situ reproduced ramets to original habitat. On the basis of the analysis of endangered mechanisms of P. armeniacum, it can be concluded that P. armeniacum has strong capability of both asexual and sexual reproductions, and an emergency mechanism consisted of massive production of rhizomes to cope with damage. Because highly effective pollinating insects that facilitate pollination in P. armeniacum are present in the habitat, flowering ramets produce fruits with large quantity of seeds, many of which in turn grow into new genets that can reproduce many cloned ramets. P. armeniacum makes very effective use of its environment and has distinct characteristics of enduring harsh environmental conditions; therefore, rather than its own inherent biological defects, the main threats facing this species are the destruction of its survival space and the wipe-out collecting of the plants as a result of trading. Based on the analysis mentioned above, certain appropriate strategies have been proposed for the conservation of P. armeniacum.     

兰科紫纹兜兰的保育生物学研究

深圳市梧桐山是濒危物种紫纹兜兰(Paphiopedilum purpuratum)仅存的几个自然分布地之一。自1998年10月1日至2002年2月28日,我们在梧桐山选取了9个调查点,对407个基株进行了观察,研究了该地的气候、植被、土壤和其他环境因素与紫纹兜兰之间的关系。观察结果表明,梧桐山地区的紫纹兜兰在次生阔叶林下生长良好;短刺刺腿食蚜蝇(Ischiodon scutellaris)为其有效传粉者;大多数开花的紫纹兜兰均能结果。我们认为,紫纹兜兰生存所面临的主要威胁应该是其生境亦即森林的破坏,而不是其自身存在的生物学缺陷。     

麻栗坡兜兰及其近缘植物的分类研究

对麻栗坡兜兰Paphiopedilum malipoense及其近亲种作了对比研究。它们被作为一个种之下的4个变种：麻栗坡兜兰（原变种）Paph.malipoense var.malipoense,浅斑兜兰（变种）var.jackii,钩唇兜兰（变种）var.hiepii;窄瓣兜兰（变种）var.angustatum（变种,新等级）。其鉴别特征已列入表中,还附有一幅图版。