published_or_final_version / Zoology / Master / Master of Philosophy

Disa comprises 163 species, 131 of which occur in South Africa (SA). The genus is distributed across winter- and summer-rainfall areas, but few species transverse both climatic regions. Species are therefore regarded as winter-rainfall or summer-rainfall endemics - yet release their seeds in autumn, irrespective of provenance. Disa contributes 40 % of threatened Orchidaceae in SA, with half of the local species requiring conservation initiatives. In vitro seed germination is a potential conservation tool for producing large numbers of genetically diverse plants in relatively short periods. However, only 11 winter-rainfall Disa species are easily germinated ex situ. Studies were therefore undertaken on summer-rainfall taxa, which are ungerminated in vitro, in an effort to define their germination parameters. This thesis describes mechanisms that control germination in Disa and establishes practical propagation methods for seed culture. Two seed types occur in Disa; i) comparatively large, pale and pyriform seeds in members of the D. uniflora sub-c1ade, which populate streamside habitats under conditions of winter-rainfall maxima, and ii) smaller, variously brown and fusiform seeds in the remainder of the genus. Seed morphometrics distinguished seed types, although embryo dimensions were similar. Testa continuity, which is disrupted in the large seeds, also supported separation. Typically, small seeds are ungerminated in vitro, whilst large seeds germinated readily. Increased seed size did not necessarily impart increased germ inability, as several germinable, small-seeded species exist - being winter-rainfall species Attempts to establish in vitro germinability revealed that increased water availability and charcoal supplementation promoted germination in intractable species. The control of germination was therefore proposed as a trade-off between water availability and the presence of phyto-inhibitors - two features typical of seeds exhibiting water-impermeable dormancy. Three germinability categories were recognized; i) easily germinable species, ii) poorly germinable species through media manipulation, and iii) ungerminated species. Germination of immature seed in the absence of media modification was comparable to mature seed germination under modified conditions, providing evidence of the role of an impermeable seed testa in regulating germination. Testa impermeability in mature, small-seeded species was demonstrated using aqueous EVANS' blue dye and was linked to i) testa integrity and ii) increased levels of leachable phenolics (LPC) - which are hydrophobic and phytotoxic. In addition, this research revealed an impervious and elaborate embryo carapace in small seeds. Large-seeded species were highly permeable at dehiscence, with perforated testae and negligible LPC. Germinability was ultimately defined by a significant regression with LPC. Phenolic deposition increased exponentially with increasing seed maturity and reflected decreased permeability and the development of testa colouration. The testa precludes the use of viability stains such as nc and FDA, unless rendered permeable through scarification. This was achieved using NaOCI. Viability and germinability percentages did not correlate well for the small-seeded Disa species, indicating that i) the methods used to break dormancy are inadequate, ii) additional factors may be acting in concert with the testa to regulate germination and iii) that the determination of mature Disa seed viability is ineffective. As an alternative, the germination potential of immature seed was estimated as the ratio between the proportion of embryos stained with TTC and the proportion of seeds permeable to EVANS' blue. Attempts to relieve water-impermeable dormancy in Disa resulted in the formulation of a dual-phase protocol - with the specific aim of increasing water availability to the embryo. Dual-phase cultures comprised a solid, charcoal-rich medium overlaid with a reduced strength, liquid medium fraction of the same type. The solid fraction negated the influence of leached phenols and allowed protocorms to establish polarity, whilst the fluid fraction increased water availability. The dual-phase protocol allowed germination of nine summer-rainfall Disa species, usually in percentages that approximated their estimated germination potential. For the remaining species, germination is controlled by more complex factors. Large seeds are atypical in containing starch, the hydrolysis of which facilitated their rapid, autonomous germination. Small-seeded Disa species stored lipids and proteins and germinable species accumulated starch post-germination. The embryo protoplasts of all species contained appreciable amounts of soluble sugars, irrespective of germinability. However, decreased sucrose and increased fructose correlated significantly with decreased seed germinability. This study provides evidence of the nutritional value of mycotrophy, with endophytes liberating soluble carbohydrate and non-carbohydrate compounds upon lysis. However, few species were germinated symbiotically, suggesting that endophytes isolated from adult roots do not necessarily support germination in the same species. Similar endophytic fungi occur in Australian and Holarctic orchids. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2003.

Odontoglossum ringspot virus (ORSV) is one of the most prevalent orchid viruses that infects greenhouse-grown orchids worldwide. In order to prevent the spread of viruses in greenhouses and to cultivate clones from virus-free mother plants, it is necessary to develop a more sensitive technique for the detection of viruses in orchids. SYBR green real-time RT-PCR is a highly sensitive technique that can specifically detect ORSV in orchid tissue. By harvesting tissue at the inoculation site and at specific distances from the inoculation site at different times past inoculation, this technique can also be used to study the rate of spread of ORSV in orchids. Orchid clones were inoculated with ORSV and other clones were mock-inoculated with molecular grade water. Leaf tissue was harvested from the ORSV-inoculated and mock-inoculated clones at the site of inoculation and at specific distances from this site at 16 h, 24 h, and 72 h past inoculation. Total RNA was extracted from the harvested tissue. Competitive RTPCR was going to be used for the quantification and detection of ORSV in the samples, but attempts at cloning an ORSV fragment into a vector in order to form a competitive standard were unsuccessful. Instead, a highly sensitive qualitative approach called SYBR green real-time RT-PCR was used for the detection of ORSV. ORSV was detected in all virus-inoculated orchids, except for one. Therefore, all of the ORSV inoculated plants except for one were infected with the virus. Unexpectedly, ORSV was also detected in all of the mock-inoculated orchids. Most likely the orchids were previously infected with ORSV, but the viral titer was too low to be detected by commercial techniques. However, there is a small possibility that the orchids were contaminated during experimentation, despite careful technique. The rate of spread of the virus could not be studied because the mock-inoculated samples also contained the virus. Although viral amplification was demonstrated in the mock-inoculated plants, SYBR green real-time RT-PCR is still a sensitive and consistent method for ORSV detection in orchids. With additional controls, this method could prove to be the ideal method for reliable detection of ORSV in commercially-grown orchids. / Department of Biology

Due to the rapid destruction of native orchid habitats it has become necessary to house many endangered orchid species in greenhouse environments where enhanced spread of viral disease occurs due to the close contact between plants. This research was concerned with the construction of a library of genes whose expression is induced in response to viral challenge. In uncovering the genes that are activated during plant-pathogen interactions, it may be possible to manipulate these pathways to develop virus resistant orchids. Furthermore, this research will contribute additional information for the existing framework of plant-pathogen interactions of all plant species.In order to construct a library of genes expressed in response to viral infection, suppression subtractive hybridization was performed using the PCR-Select cDNA Subtraction Kit (CLONTECH, Palo Alto, CA) on Sophrolaeliacattleya Ginny Champion 'Riverbend' clones. RNA was isolated from plants that had been inoculated with the Odontoglossum ringspot virus (ORSV) and from control plants that had not been inoculated with ORSV. Following reverse transcription-PCR (RT-PCR) to obtain cDNA, cDNAs of the tester population (those cDNAs containing differentially expressed messages in response to ORSV) and the driver population (reference cDNAs from uninfected plants) were obtained. The two different cDNA populations are mixed together and hybridized. The sequences common to both populations were subtracted, leaving only the differentially expressed sequences available for PCR amplification.A library containing these genes was constructed, and one clone, chosen at random, was sequenced. Based on homology comparisons to known genes, we have cloned a gene that may contain a nucleotide binding site similar to that of the tobacco N gene, important for plant resistance to pathogens. In the near future, this clone will be used to construct probes for use in northern analysis to determine the timing and localization of the products of this gene. This information will aid in characterizing the function of the orchid N-gene and identifying other members of this signal cascade. In addition, many other clones await sequencing and similar characterization. / Department of Biology

Thesis (M.S.)--University of Hawaii at Manoa, 2007. / Includes bibliographical references (leaves 98-110). / ix, 110 leaves, bound ill. (some col.) 29 cm

Orientador: Kathia Fernandes Lopes Pivetta / Coorientador: Ricardo Tadeu de Faria / Coorientador: Wagner Aparecido Vendrame / Banca: Renata Gimenes / Banca: Marcos Vieira Ferraz / Banca: Paulo Hercilio Viegas Rodrigues / Banca: Claudia Fabrino Machado Mattiuz / Resumo: As plantas da família Orchidaceae são muito apreciadas pelo potencial ornamental, ecológico e econômico. O domínio de técnicas para a domesticação e propagação em massa das espécies é extremamente importante, visto que, possibilita diminuir a coleta predatória, além de reduzir o custo de produção das plantas. O cultivo in vitro é uma técnica que permite produzir grande número de plantas; entretanto, ocorrem muitas perdas durante o período de aclimatização (ex vitro). O aprimoramento dos métodos de conservação de recursos genéticos por meio de criopreservação de sementes em desenvolvimento é uma importante estratégia para a conservação de germoplasma e programas de melhoramento genético de plantas desta família. Este trabalho teve como objetivos avaliar o desenvolvimento in vitro de plântulas em meio de cultura alternativo e diferentes ambientes das orquídeas Amblostoma amblostomoides e Cattleya percivaliana, bem como, diferentes protocolos para a criopreservação de sementes da orquídea Ionopsis utricularioides. Nos experimentos sobre desenvolvimento in vitro de plântulas em meio de cultura alternativo e diferentes ambientes, para cada espécie, o delineamento experimental utilizado foi o inteiramente casualizado. Inicialmente foram seis tratamentos (T1 - meio MS (Murashige e Skoog) + sala de crescimento - Controle; T2 - MCA (meio de cultura alternativo) + sala de crescimento; T3 - MCA + tela preta com 70% de sombreamento; T4 - MCA + tela azul com 50% de sombreamento; T5 - MCA + tela preta com 50% de sombreamento e T6 - MCA + tela vermelha com 50% de sombreamento) e quatro repetições com 10 plântulas por parcela avaliando-se porcentagem de sobrevivência; para avaliação dos dados biométricos, em razão do alto índice de mortalidade, foram avaliados quatro tratamentos para A. amblostomoides e três tratamentos para C. percivaliana, com 12 repetições para ambas as espécies, sendo uma plântula... / Abstract: Orchids are well appreciated by ornamental, ecological and economic potential. The improvement of genetic resources conservation methods through seeds and protocorms cryopreservation is an important strategy for the conservation of germplasm and plant breeding programs of this family. The domain of techniques for domestication and mass propagation of species is very important because, enables decreasing predation and reduce the cost of production of plants. The in vitro culture is a technique that allows the production of a large numbers of plants; however, there are many losses during the acclimatization period (ex vitro). This work aimed to evaluate different protocols for cryopreservation of Ionopsis utricularioides seeds, as well as the in vitro development of seedlings in alternative culture media and different environments of the orchids, Amblostoma amblostomoides and Cattleya percivaliana. In experiments on cryopreservation of seeds for each species, the experimental design was completely randomized with eight treatments and four replications each. The treatments were: T1) seeds without cryopreservation (control 1); T2) seeds cryopreserved without cryoprotectants (Control 2); T3) glycerol + PVS2; T4) glycerol + PVS2 + phloroglucinol; T5) sucrose + PVS2; T6) sucrose + PVS2 + phloroglucinol; T7) glycerol + sucrose + PVS2; and T8) + sucrose + glycerol + PVS2 + phloroglucinol and 10 repetitions. The variables analyzed were germination percentage, number of seedlings and protocorms and development of the seedlings formed. In the experiments on in vitro development of seedlings in an alternative culture media and in different environments, the experimental design was completely randomized for each species. There were six treatments [T1: MS medium (Murashige e Skoog) - under laboratory conditions; T2: ACM (Alternative Culture Medium) - under laboratory conditions; T3: MCA - black net with 50% shading; T4: MCA - black net with ... / Doutor

Made available in DSpace on 2016-04-01T17:55:03Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-08-31. Added 1 bitstream(s) on 2016-04-01T18:00:53Z : No. of bitstreams: 1 000859876_20160630.pdf: 471050 bytes, checksum: b283e5cdd6fee1565d98e89bf93e40b0 (MD5) Bitstreams deleted on 2016-07-01T13:02:23Z: 000859876_20160630.pdf,. Added 1 bitstream(s) on 2016-07-01T13:03:18Z : No. of bitstreams: 1 000859876.pdf: 729708 bytes, checksum: 0c3f9644fb91e37bed700362170aae04 (MD5) / As plantas da família Orchidaceae são muito apreciadas pelo potencial ornamental, ecológico e econômico. O domínio de técnicas para a domesticação e propagação em massa das espécies é extremamente importante, visto que, possibilita diminuir a coleta predatória, além de reduzir o custo de produção das plantas. O cultivo in vitro é uma técnica que permite produzir grande número de plantas; entretanto, ocorrem muitas perdas durante o período de aclimatização (ex vitro). O aprimoramento dos métodos de conservação de recursos genéticos por meio de criopreservação de sementes em desenvolvimento é uma importante estratégia para a conservação de germoplasma e programas de melhoramento genético de plantas desta família. Este trabalho teve como objetivos avaliar o desenvolvimento in vitro de plântulas em meio de cultura alternativo e diferentes ambientes das orquídeas Amblostoma amblostomoides e Cattleya percivaliana, bem como, diferentes protocolos para a criopreservação de sementes da orquídea Ionopsis utricularioides. Nos experimentos sobre desenvolvimento in vitro de plântulas em meio de cultura alternativo e diferentes ambientes, para cada espécie, o delineamento experimental utilizado foi o inteiramente casualizado. Inicialmente foram seis tratamentos (T1 - meio MS (Murashige e Skoog) + sala de crescimento - Controle; T2 - MCA (meio de cultura alternativo) + sala de crescimento; T3 - MCA + tela preta com 70% de sombreamento; T4 - MCA + tela azul com 50% de sombreamento; T5 - MCA + tela preta com 50% de sombreamento e T6 - MCA + tela vermelha com 50% de sombreamento) e quatro repetições com 10 plântulas por parcela avaliando-se porcentagem de sobrevivência; para avaliação dos dados biométricos, em razão do alto índice de mortalidade, foram avaliados quatro tratamentos para A. amblostomoides e três tratamentos para C. percivaliana, com 12 repetições para ambas as espécies, sendo uma plântula... / Orchids are well appreciated by ornamental, ecological and economic potential. The improvement of genetic resources conservation methods through seeds and protocorms cryopreservation is an important strategy for the conservation of germplasm and plant breeding programs of this family. The domain of techniques for domestication and mass propagation of species is very important because, enables decreasing predation and reduce the cost of production of plants. The in vitro culture is a technique that allows the production of a large numbers of plants; however, there are many losses during the acclimatization period (ex vitro). This work aimed to evaluate different protocols for cryopreservation of Ionopsis utricularioides seeds, as well as the in vitro development of seedlings in alternative culture media and different environments of the orchids, Amblostoma amblostomoides and Cattleya percivaliana. In experiments on cryopreservation of seeds for each species, the experimental design was completely randomized with eight treatments and four replications each. The treatments were: T1) seeds without cryopreservation (control 1); T2) seeds cryopreserved without cryoprotectants (Control 2); T3) glycerol + PVS2; T4) glycerol + PVS2 + phloroglucinol; T5) sucrose + PVS2; T6) sucrose + PVS2 + phloroglucinol; T7) glycerol + sucrose + PVS2; and T8) + sucrose + glycerol + PVS2 + phloroglucinol and 10 repetitions. The variables analyzed were germination percentage, number of seedlings and protocorms and development of the seedlings formed. In the experiments on in vitro development of seedlings in an alternative culture media and in different environments, the experimental design was completely randomized for each species. There were six treatments [T1: MS medium (Murashige e Skoog) - under laboratory conditions; T2: ACM (Alternative Culture Medium) - under laboratory conditions; T3: MCA - black net with 50% shading; T4: MCA - black net with ...

Made available in DSpace on 2014-06-11T19:23:02Z (GMT). No. of bitstreams: 0 Previous issue date: 2013-04-09Bitstream added on 2014-06-13T19:08:48Z : No. of bitstreams: 1 andreota_rc_me_rcla.pdf: 1095020 bytes, checksum: 849f970f9b61ff625a1f282da278410b (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / A família Orchidaceae compreende cerca de 25.000 espécies distribuídas em cinco subfamílias. Os representantes terrícolas constituem cerca de 20% das orquídeas e encontramse incluídos principalmente na subfamília Orchidoideae, dentro da qual se encontra a tribo Cranichideae. No Brasil a tribo Cranichideae possui representantes das subtribos Cranichidinae, Goodyerinae e Spiranthinae. No presente estudo foram analisados os órgãos vegetativos das seguintes espécies: Cranichis candida e Prescottia oligantha (Cranichidinae), Microchilus arietinus e Zeuxine strateumatica (Goodyerinae), Cyclopogon apricus, C. congestus, C. variegatus, Mesadenella cuspidata, Pteroglossa roseoalba e Sauroglossum nitidum (Spiranthinae), visando apontar caracteres estruturais relacionados ao hábito terrestre e potencialmente úteis na delimitação taxonômica do grupo. As raízes estudadas constituem, proporcionalmente, o órgão mais desenvolvido da planta e apresentam características que podem auxiliar na absorção/retenção de água, como a ocorrência de um sistema velame/exoderme, bem como a presença de tilossomos nas Cranichidinae e Spiranthinae. As folhas são portadoras de mesofilos estreitos e homogêneos, recobertas por epiderme unisseriada e revestidas por cutículas finas. Os feixes condutores da nervura central são envolvidos por células parenquimáticas e as células do xilema se dispõem ao pares ou em forma de V invertido, como nas demais orquídeas da tribo Cranichideae / Orchidaceae comprises ca. 25.000 species currently divided in five subfamilies. Terrestrial species constitute almost 20% of the family and are included mainly in the subfamily Orchidoideae, to which belong the tribe Cranichideae. In Brazil Cranichideae is represented by species of subtribes Cranichidinae, Goodyerinae and Spiranthinae. In this study, vegetative organs of the following species were analised: Cranichis candida and Prescottia oligantha (Cranichidinae), Microchilus arietinus and Zeuxine strateumatica (Goodyerinae), Cyclopogon apricus, C. congestus, C. variegatus, Mesadenella cuspidata, Pteroglossa roseoalba and Sauroglossum nitidum (Spiranthinae). The aims were to find structural features related to the terrestrial habit and potentially useful in the taxonomic delimitation of the group. The roots are, proportionally, the most developed organ of the plant, showing characteristics that might help in absorbing or retaining water, such as: velamen/exoderm system, in all species, and tilossomes, in the Cranichidinae and Spiranthinae representatives. Leaves possess an one-layered epidermis covered by a thin cuticle and a narrow and homogeneous mesophyll. Vascular bundles of the midrib are enclosed by parenchymatic cells and the xylem conducting cells are organised in pairs or “inverted V”, as in other Cranichideae

Orientador: Maria das Graças Sajo / Coorientador: Fábio de Barros / Banca: Sandra Maria Carmello Guerreiro / Banca: Adelita Aparecida Sartori Paoli / Resumo: A família Orchidaceae compreende cerca de 25.000 espécies distribuídas em cinco subfamílias. Os representantes terrícolas constituem cerca de 20% das orquídeas e encontramse incluídos principalmente na subfamília Orchidoideae, dentro da qual se encontra a tribo Cranichideae. No Brasil a tribo Cranichideae possui representantes das subtribos Cranichidinae, Goodyerinae e Spiranthinae. No presente estudo foram analisados os órgãos vegetativos das seguintes espécies: Cranichis candida e Prescottia oligantha (Cranichidinae), Microchilus arietinus e Zeuxine strateumatica (Goodyerinae), Cyclopogon apricus, C. congestus, C. variegatus, Mesadenella cuspidata, Pteroglossa roseoalba e Sauroglossum nitidum (Spiranthinae), visando apontar caracteres estruturais relacionados ao hábito terrestre e potencialmente úteis na delimitação taxonômica do grupo. As raízes estudadas constituem, proporcionalmente, o órgão mais desenvolvido da planta e apresentam características que podem auxiliar na absorção/retenção de água, como a ocorrência de um sistema velame/exoderme, bem como a presença de tilossomos nas Cranichidinae e Spiranthinae. As folhas são portadoras de mesofilos estreitos e homogêneos, recobertas por epiderme unisseriada e revestidas por cutículas finas. Os feixes condutores da nervura central são envolvidos por células parenquimáticas e as células do xilema se dispõem ao pares ou em forma de V invertido, como nas demais orquídeas da tribo Cranichideae / Abstract: Orchidaceae comprises ca. 25.000 species currently divided in five subfamilies. Terrestrial species constitute almost 20% of the family and are included mainly in the subfamily Orchidoideae, to which belong the tribe Cranichideae. In Brazil Cranichideae is represented by species of subtribes Cranichidinae, Goodyerinae and Spiranthinae. In this study, vegetative organs of the following species were analised: Cranichis candida and Prescottia oligantha (Cranichidinae), Microchilus arietinus and Zeuxine strateumatica (Goodyerinae), Cyclopogon apricus, C. congestus, C. variegatus, Mesadenella cuspidata, Pteroglossa roseoalba and Sauroglossum nitidum (Spiranthinae). The aims were to find structural features related to the terrestrial habit and potentially useful in the taxonomic delimitation of the group. The roots are, proportionally, the most developed organ of the plant, showing characteristics that might help in absorbing or retaining water, such as: velamen/exoderm system, in all species, and tilossomes, in the Cranichidinae and Spiranthinae representatives. Leaves possess an one-layered epidermis covered by a thin cuticle and a narrow and homogeneous mesophyll. Vascular bundles of the midrib are enclosed by parenchymatic cells and the xylem conducting cells are organised in pairs or "inverted V", as in other Cranichideae / Mestre

Submitted by Liliane Ferreira (ljuvencia30@gmail.com) on 2018-11-30T13:53:38Z No. of bitstreams: 2 Dissertação - Daniella Mota Silva - 2012.pdf: 3271983 bytes, checksum: 99a6d63c282c21b5c3ced5fe6efe4944 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Rejected by Luciana Ferreira (lucgeral@gmail.com), reason: O resumo em português está cortado, olhe no formulário de metadados se está completo. on 2018-12-03T13:12:51Z (GMT) / Submitted by Liliane Ferreira (ljuvencia30@gmail.com) on 2018-12-04T13:07:29Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação - Daniella Mota Silva - 2012.pdf: 3271983 bytes, checksum: 99a6d63c282c21b5c3ced5fe6efe4944 (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2018-12-05T09:55:21Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação - Daniella Mota Silva - 2012.pdf: 3271983 bytes, checksum: 99a6d63c282c21b5c3ced5fe6efe4944 (MD5) / Made available in DSpace on 2018-12-05T09:55:21Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação - Daniella Mota Silva - 2012.pdf: 3271983 bytes, checksum: 99a6d63c282c21b5c3ced5fe6efe4944 (MD5) Previous issue date: 2012-10-30 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / CyrtopodiumsaintlegerianumRchb. f is an epiphytic species typical of the Midwest, especially in distributed Brazilian Central Plateau, and has a wide geographical distribution in Brazil. It is usually found in the trunks of palm trees, forming large clumps. It has good features for ornamentation, for the beauty and size of its inflorescence, however, are not found in the literature about its conservation, methods for its spread or that could be used in floriculture and landscaping. Thus, this study aimed to establishing protocols for germination asymbiotic effects of phytohormones and acclimatization and characterization of chromosomal species. In 2010, the establishment of micropropagation protocols, capsules were collected in a pasture area in the municipality of Mossâmedes, GO, then the part previously sterilized seeds were separated and tested in a 1% tetrazolium dye.. For cultivation was tested asymbiotic different culture media and then tested after different concentrations and combinations in treatments BAP 16 / ANA, to finally evaluate the acclimatization substrates combined and fertilized with chemical fertilizers and organic. For the karyotype of the species, the plant material is derived from plants grown in vitro in culture medium. We tested four protocols, with differences in enzyme solution for softening the roots, dyes, anti-mitotic concentration of the solution and hydrolysis, and the use of growth regulators for root induction in vitro. All protocols were in common roots pretreated with anti-mitotic 8-hydroxyquinoline (0.002 M) in refrigerator for 24 hours. Then protocols roots were fixed in Carnoy 3:1 for 18 hours the first and second and third and fourth protocol for 24 hours at room temperature. After stored at -20 º C in the same fixative for further analysis (only the fourth protocol). The roots of the protocols were stained with different dyes: hematoxylin, Schiff, acetic orcein and Giemsarespectively.The results of germination was satisfactory in all culture media. For medium supplemented with auxin / cytokinin combined the best concentrations for variable height were 0.2 mg L-1 NAA and BAP without adding control without addition of regulators. The best means to induce large numbers of shoots were 4 mg L-1 BAP and 4 mg L-1 BAP / 0.2 mg L-1 NAA. The number of leaves was rated best in the concentrations of BAP without NAA at concentrations of 1.0, 2.0 and 4.0 mg L-1. The treatments with the highest number of roots were control without added growth regulators at doses of 0.2, 0.5, 1.0 mg L-1 NAA without addition of BAP, as well as the length of roots was favored by the same treatments. The largest number occurred in callus treatment with concentrations of 1.0 mg L-1 BAP without adding ANA. ForcytogeneticsC. saintlegerianum the best protocol was evaluated with the regulator which was obtained metaphases, however chromosomes were condensed, and the number of chromosomes was found to be 2n = 48. cides. / CyrtopodiumsaintlegerianumRchb. f é espécie epífita da região Centro-Oeste, distribuída no Planalto Central brasileiro, e tem ampla distribuição geográfica no Brasil. É encontrada em de troncos de palmeiras, formando grandes touceiras. Possui características para ornamentação, pela beleza de sua inflorescência, contudo não são encontrados na literatura estudos sobre sua conservação ou propagação. Assim, esse trabalho teve como objetivo o estabelecimento de protocolos para germinação assimbiótica, efeitos de fitohormônios, aclimatização e a caracterização cromossômica da espécie. Em 2010, para o estabelecimento de protocolos para a micropropagação, cápsulas foram coletadas em uma área de pastagem no município de Mossâmedes, GO, foram previamente desifestadas em seguida parte das sementes foram separadas e testadas em corante tetrazólio a 1%. Para o cultivo assimbiótico, foram testados diferentes meios de cultura e diferentes concentrações e combinações de BAP/ANA em 16 tratamentos. Foram testados substratos combinados e adubação com fertilizante químico e orgânico para a aclimatização das plântulas. Para o cariótipo da espécie, o material vegetal foi proveniente de plantas cultivadas in vitro em meio de cultura. Foram testados quatro protocolos, com diferenças quanto a solução enzimática para o amolecimento das raízes, os corantes, concentração do antimitótico e solução de hidrólise, e o uso de regulador de crescimento para indução de raízes in vitro. Todos os protocolos tiveram em comum raízespré-tratadas com anti-mitótico 8- hidroxiquinoleína (0,002M) em geladeira durante 24 horas. Em seguida nos protocolos as raízes foram fixadas em Carnoy 3:1 por 18 horas o primeiro e o segundo protocolo e o terceiro e o quarto por 24 horas em temperatura ambiente. Depois estocados a -20ºC no próprio fixador, para posterior análise (apenas o quarto protocolo). As raízes dos protocolos foram coradas com diferentes corantes: hematoxilina, reativo de Schiff, orceína acética e Giemsa respectivamente. A germinação foi satisfatória em todos os meios de cultura. Para o meio suplementado com auxina/citocinina combinadas as melhores concentrações para a variável altura foi 0,2 mg L-1 de ANA sem adição de BAP e o controle sem adição de reguladores. Os melhores meios que induziram grande número de brotações foram 4 mg L-1 de BAP e 4 mg L-1 de BAP / 0,2 mg L-1 de ANA. O número de folhas foi melhor avaliado nas concentrações de BAP sem ANA nas concentrações 1,0; 2,0 e 4,0 mg L-1. Os tratamentos com maior número de raízes foram o controle sem adição de reguladores de crescimento, e nas dosagens de 0,2, 0,5, 1,0 mg L-1 de ANA sem adição de BAP, assim como o comprimento da maior raiz foi favorecido pelos mesmos tratamentos. O maior número de calos se deu no tratamento com concentrações de 1,0 BAP mg L-1 sem adição de ANA. Para a citogenética de C. saintlegerianumo melhor protocolo avaliado foi com regulador no qual foi obtido metáfases, no entanto os cromossomos se encontravam condensados, e o numero de cromossomos encontrado foi de 2n=48.