Development of the Embryo Sac in Amitostigma kinoshitae (Makino) Schltr. (Orchidaceae)

The embryo sac of Amitostigma kinoshitae was studied. The ovuleis anatropous, bitegmic, and tenuinucellate. The inner integumentalone forms a micropyle. The megaspore of a tetrad nearest tothe chalaza develops into an eight-nucleate embryo sac of thetypical Polygonum-type. Double fertilization takes place normally.     

Gastrodia elata Blume (tianma) mobilizes neuro-protective capacities

Tianma (Gastrodia elata Blume) is a traditional Chinese medicine (TCM) often used for the treatment of headache, convulsions, hypertension and neurodegenerative diseases. Tianma also modulates the cleavage of the amyloid precursor protein App and cognitive functions in mice. The neuronal actions of tianma thus led us to investigate its specific effects on neuronal signalling. Accordingly, this pilot study was designed to examine the effects of tianma on the proteome metabolism in differentiated mouse neuronal N2a cells using an iTRAQ (isobaric tags for relative and absolute quantitation)-based proteomics research approach. We identified 2178 proteins, out of which 74 were found to be altered upon tianma treatment in differentiated mouse neuronal N2a cells. Based on the observed data obtained, we hypothesize that tianma could promote neuro-regenerative processes by inhibiting stress-related proteins and mobilizing neuroprotective genes such as Nxn, Dbnl, Mobkl3, Clic4, Mki67 and Bax with various regenerative modalities and capacities related to neuro-synaptic plasticity.     

鹤顶兰胚囊发育过程中Ca~（2＋）分布的超微细胞化学定位（英文）

用焦锑酸盐沉淀法对鹤顶兰（Phaius tankervilliae）胚囊发育过程中的Ca2＋状态进行超微细胞化学定位。观察结果发现：功能大孢子时期,珠孔端的胚囊壁上开始出现小颗粒的Ca2＋沉淀,但功能大孢子细胞内未见明显的Ca2＋标记;四核胚囊时期胚囊壁上的Ca2＋沉淀明显增多,液泡膜上有Ca2＋沉淀出现,珠孔处的Ca2＋沉淀颗粒较大;成熟胚囊时期,胚囊壁上的Ca2＋沉淀进一步增多,且胚囊内Ca2＋分布明显增多,且极性明显,珠孔端助细胞、卵细胞比合点端反足细胞有更多的Ca2＋沉淀。鹤顶兰成熟胚囊内Ca2＋积累的来源有：（1）在胚囊成熟前主要由珠被细胞、珠细胞通过胞间连丝向胚囊运输;（2）以沉淀有大量Ca2＋的小泡形式跨过胚囊壁进入胚囊。     

天麻中天麻素含量的影响因子研究

应用高压液相色谱技术,对产地、品种、以及炮制方法等因素对天麻中天麻素含量的影响进行了比较分析.结果表明,炮制对天麻药材中天麻素的含量影响最大.对不同炮制方法的比较分析结果显示,蒸制法可显著提高天麻素的含量.本文还讨论了天麻素在炮制过程中形成的可能机理.     

天麻成分对羟基苯甲醛抗血小板聚集作用及急性毒性研究

为探讨天麻成分对羟基苯甲醛的抗血小板聚集作用,本实验以二磷酸腺苷(ADP)、花生四烯酸(AA)、血小板活化因子(PAF)为诱导剂,探讨了对羟基苯甲醛的体外抗血小板聚集活性;采用ADP静脉注射致小鼠肺栓塞方法以及下腔静脉结扎致大鼠静脉血栓方法,考察了对羟基苯甲醛的体内抗血小板聚集活性;并对经口给药的对羟基苯甲醛的急性毒性进行了检测。结果表明,天麻成分对羟基苯甲醛对ADP诱导的家兔体外血小板聚集有明显的对抗作用,其半数抑制率(50%inhibitory concentration,IC50)为2mmol/L,对PAF诱导的家兔体外血小板聚集无明显影响;天麻成分对羟基苯甲醛能明显降低小鼠肺栓塞死亡率,并显著对抗大鼠下腔静脉血栓的形成。说明天麻成分对羟基苯甲醛在体外、体内均具有显著的抗血小板聚集活性。急性毒性研究结果表明天麻成分对羟基苯甲醛经口给药的小鼠半数致死量(lethal dose 50,LD50)为1.23 g/kg。     

天麻酯溶性酚性成分对大鼠脑缺血/再灌注损伤的保护作用

为探讨天麻酯溶性酚性成分对大鼠脑缺血/再灌注的保护作用,本实验采用线栓法复制了大鼠局灶性脑缺血再灌注损伤模型,以神经学评分、脑梗塞体积、脑组织内水分含量、脑组织内钙调蛋白依赖性蛋白激酶(CaMKⅡ)表达作为评价指标,考察天麻酯溶性酚性成分的作用,结果表明天麻酯溶性酚性成分能够明显改善模型动物的神经病学症状,缩小脑梗塞体积,减轻脑水肿,调节CaMKⅡ的表达水平。说明天麻酯溶性酚性成分在防治大鼠脑缺血/再灌注损伤方面具有明显作用。     

天麻属(兰科)一新变种——卵果天麻

记载了陕西天麻属一新变种--卵果天麻(Gastrodia elata var.obovata Y.J.Zhang).新变种与原变种不同在于花螺旋排列,果实短圆锥型偏斜,花茎多为3节.     

黔产天麻的化学成分

从黔产天麻（Ｇａｓｔｒｏｄｉａ ｅｌａｔａ Ｂｌ．）中分离得到１个新的含氮化合物,命名为天麻羟胺（ｇａｓｔｒｏｄａｍｉｎｅ,Ⅰ）,经波谱分析,确定其结构为双－（对羟苄基）羟胺。除β－谷甾醇和胡萝卜甙外,同时还分离鉴定了３个已知化合的：Ｌ－焦谷氨酸（Ⅱ）、柠檬酸单甲酯（Ⅲ）、柠檬酸双甲酯（Ⅳ）,其中化合物Ⅱ和Ⅳ为首次报道从天麻中得到的。     

乌天麻Gastrodianin基因的鉴定及其在天麻发育进程中的表达

基于相似性克隆策略和RT-PCR,从兰科植物乌天麻(Gastrodia elataf.glauca)球茎克隆得到两个新的天麻抗真菌蛋白基因,命名为gastrodianin-4A(ga4A)和gastrodianin-4B(ga4B),并首次用Northern杂交研究了该基因在植物不同部位的转录表达。结果表明,天麻个体的不同部位只转录表达同一种Gastrodianin基因,纯化自球茎的Gastrodianin蛋白质的肽质量指纹谱与推导的Ga4A和Ga4B成熟蛋白的氨基酸序列相吻合;Northern杂交证明天麻的地上器官Gastrodianin基因转录表达量远远高于地下球茎,而次生球茎皮层组织的表达量比中柱和整个营繁茎的都高一些。天麻球茎Gastrodianin的外周表达模式可能是天麻在地下抵御蜜环菌(Armillaria mellea Karst)入侵球茎皮层内部的防卫机制之一,但该基因在天麻地上部分的高丰度表达暗示Gastrodianin可能蕴藏其它生理功能。     

Gastrodia sui sp. nov. (Orchidaceae) from Taiwan

A new orchid species, Gastrodia sui C. S. Leou, T. C. Hsu & C. L. Yeh is described and illustrated from Taiwan. The new species is closely related to G. callosa J. J. Sm. but distinguishable by the number and position of calli inside the perianth tube, the fused ratio of perianth lobes, and the structure of the lip and column. It is so far only found in the area of Mt Lilung in southern Taiwan.     

莴苣胚囊细胞分离

用酶解和解剖方法分离了莴苣的卵细胞,助细胞,中央细胞和合子。莴苣子房先在酶液中酶解40～50min,然后在不含酶的分离液中用解剖针解剖子房。在解剖出的胚囊中,可看到卵细胞,两个助细胞和中央细胞的轮廓。将胚囊的合点端切破,轻轻挤压胚囊的珠孔端,四个细胞即可逸出。在最佳条件下,90min可从40个子房中分离出29个胚囊,进一步从中分离出11个卵细胞。分离出的胚囊细胞用显微操作仪收集备用。莴苣卵细胞的成功分离为进行离体受精探索创造了条件。     

Two Types of Embryo Sac Development in Scrophularia himalensis Royle

The development of the embryo sac in Scrophularia himalensisRoyle (tribe Cheloneae, Scrophulariaceae) has been investigated.The ovules are anatropous, unitegminal and tenuinucellar. Twotypes of embryo sac development—the monosporic (polygonumtype) and the bisporic (allium type)—coexist in this species.This is the first record of its kind in Scrophularia as previousworkers reported only the Polygonum type of development in thetribe Cheloneae.     

Molecular phylogeny of Phalaenopsis Blume (Orchidaceae) on the basis of plastid and nuclear DNA

Representatives of the genus Ancistrocladus, woody lianas of tropical Africa and Asia, contain pharmaceutically interesting alkaloids and have been the subjects of intensive phytochemical investigations. In Southeast Asia, Ancistrocladus tectorius, previously regarded as the only species of the genus from this region, is extremely polymorphic with respect to naphthylisoquinoline alkaloids, indicating that this taxon might be comprised of several morphologically similar species. We performed a comparative study of the ITS region of nuclear rDNA and of the trnK intron of cpDNA as well as an ISSR fingerprint analysis. Using 75 samples of A. tectorius from 21 locations in comparison to samples of other Ancistrocladus species from Asia and from West and Central Africa, we investigated patterns of species differentiation within this taxon. We found the high variability of chemical compounds described for A. tectorius to be paralleled by a high genetic variability of the units that have been assigned to this taxon. Samples assigned to A. tectorius were paraphyletic with respect to species from the Indian subcontinent, and intraspecific variability was comparable to interspecific variability among the African taxa. In addition, groups of individuals occurring in sympatry were found to be more similar to those from other locations, suggesting low levels of gene flow between those sympatric groups. This indicates either a considerable number of hybridization events during the evolution of A. tectorius or the existence of several distinguishable species not yet recognized. Our results are a first step in the development of species or population-specific markers for the prediction of the alkaloid spectrum of samples. This will help to improve the reproducibility of phytochemical research on Ancistrocladus.     

天麻内生真菌菌群组成的初步研究

从采自贵州、陕西、四川和云南的天麻组织中共分离获得62株内生真菌。经形态学与分子生物学相结合鉴定,62株内生真菌分属于3个亚门、5纲、7目、9科、13属。研究结果还表明采自不同地点的天麻以及天麻不同部位的内生真菌的类群与分布存在明显差异。     

非洲狼尾草未受精无孢子生殖胚囊退化研究

为理解植物无孢子生殖胚囊未受精条件下的退化,对无孢子生殖植物非洲狼尾草未受精成熟胚囊中央细胞退化做了细胞形态学研究。没有受精的中央细胞退化时最显著的特点是细胞核产生核膜囊泡。核膜囊泡有两种类型:单层膜的囊泡和双层膜的囊泡,单层膜囊泡在细胞质中,双层膜囊泡在细胞核内。核膜囊泡有两种发生方式:1)核膜的外膜向细胞质一侧膨胀产生囊泡,囊泡进入细胞质;2)核膜向核内凹陷形成囊泡,囊泡进入细胞核。核膜囊泡类型与产生方式密切关联。核膜囊泡吞噬并消化包括线粒体在内的细胞质和核质。     

Female Gametophyte Development in Maize: Microtubular Organization and Embryo Sac Polarity

The developmental stages of the maize embryo sac were correlated with the corresponding silk lengths of ear florets in the female inflorescence. The development of embryo sacs in the ovules of spikes occurs in a gradient pattern with the initiation of the embryo sac beginning at the base of the ear and progressing to the top. At the beginning of meiosis, the presence of conspicuous cortical microtubules coincides with the extensive elongation of the megasporocyte. The spindles at metaphase I and II align along the long axis of the megasporocyte leading to the linear alignment of the dyad and tetrad of megaspores. During megagametogenesis, micropylar and chalazal nuclei of the embryo sac undergo synchronized divisions and migration at the second and third mitosis. Radiate perinuclear microtubules are present during the interphase of the second and third mitosis, and inter-sister nuclear microtubules occur at the late four-nucleate embryo sac. The configuration and orientation of the spindles, phragmoplasts, and pairs of nuclei result in precise positioning of the nuclei. The fusion of the polar nuclei and the formation of a microtubule organizing center-like structure in the filiform apparatus occur right after the first division of the antipodal cells. The different patterns of organization of microtubules in the cells of the mature embryo sac reflect their structural adaptations for their future function.     

Molecular phylogeny of Phalaenopsis Blume (Orchidaceae) based on the internal transcribed spacer of the nuclear ribosomal DNA

The internal transcribed spacer (ITS1, 5.8S rDNA, and ITS2) region of nuclear ribosomal DNA (nrDNA) was sequenced from 53 species, which represent most of the living species diversity in the genus Phalaenopsis (Orchidaceae). A phylogeny was developed for the genus based on the neighbor-joining and maximum parsimony analyses of molecular data. Results of these analyses provided support for the monophyly of the genus Phalaenopsis and concurred in that the genera Doritis and Kingidium should be treated as being parts of the genus Phalaenopsis as suggested by Christenson (2001). Within the genus Phalaenopsis, neither subgenera Aphyllae nor Parishianae were monophyletic, and they were highly clustered with subgenus Proboscidioides plus sections Esmeralda and Deliciosae of subgenus Phalaenopsis based on ITS data. Those species also have the same characters of morphology of four pollinia and similar biogeographies. Furthermore, neither subgenus Phalaenopsis nor Polychilos was monophyletic. Within the subgenus Phalaenopsis, only section Phalaenopsis was highly supported as being monophyletic. As for the subgenus Polychilos, only section Polychilos was moderately supported as being monophyletic. In conclusion, the present molecular data obtained from the ITS sequence of nrDNA of the genus Phalaenopsis provide valuable information for elucidating the phylogeny of this genus.     

天麻rDNA ITS区的PCR-RFLP分析

对来源于贵州大方（DF）、黎平（LP）、台江（TJ）、雷山（LS）、乌当（WD）及云南（YN）的鲜品天麻进行核糖体DNA内转录间隔区（ITS）的PCR扩增,并用限制性内切酶BamHI、HincⅡ,HeaⅢ进行酶切分析的结果显示,天麻ITS区长度约为750bp,PCR产物经HeaⅢ酶切后YN、DF和WD株的RFLP图谱基上一致,TJ2、LP和Ls株的RFLP图谱基本上一致,而TJ1株则显著不同;经BamHI酶切后,LP、TJ和LS株的RFLP图谱相同,除DF3外,YN、DF1、DF2和WD株的RFLP图谱基本上一致;经HincⅡ酶切后,除DF3和TJ1 RFLP的图谱显著不同外,其余样本RFLP图谱均表现一致。表明天麻的遗传变异特征与其地域分布有一定的联系。     

Gastrodia anti-fungal protein from the orchid Gastrodia elata confers disease resistance to root pathogens in transgenic tobacco

Diseases of agricultural crops are caused by pathogens from several higher-order phylogenetic lineages including fungi, straminipila, eubacteria, and metazoa. These pathogens are commonly managed with pesticides due to the lack of broad-spectrum host resistance. Gastrodia anti-fungal protein (GAFP; gastrodianin) may provide a level of broad-spectrum resistance due to its documented anti-fungal activity in vitro and structural similarity to insecticidal lectins. We transformed tobacco (Nicotiana tabacum cv. Wisconsin 38) with GAFP-1 and challenged transformants with agriculturally important plant pathogens from several higher-order lineages including Rhizoctonia solani (fungus), Phytophthora nicotianae (straminipile), Ralstonia solanacearum (eubacterium), and Meloidogyne incognita (metazoan). Quantitative real-time PCR and western blotting analysis indicated that GAFP-1 was transcribed and translated in transgenic lines. When challenged by R. solani and P. nicotianae, GAFP-1 expressing lines had reduced symptom development and improved plant vigor compared to non-transformed and empty vector control lines. These lines also exhibited reduced root galling when challenged by M. incognita. Against R. solanacearum expression of GAFP-1 neither conferred resistance, nor exacerbated disease development. These results indicate that heterologous expression of GAFP-1 can confer enhanced resistance to a diverse set of plant pathogens and may be a good candidate gene for the development of transgenic, root-disease-resistant crops.