Morpho-anatomy of in vitro germination and cryopreservation of the orchid Cattleya crispa (Orchidaceae Morfoanatomía de la germinación in vitro y criopreservación de la orquídea Cattleya crispa (Orchidaceae) / Morfoanatomía de la germinación in vitro y criopreservación de la orquídea Cattleya crispa (Orchidaceae)

Introduction: Cattleya crispa is an ornamental epiphytic orchid with geographic distribution restricted to the Brazilian Atlantic Forest. Due to predatory extractivism and human-induced habitat loss, this species appears on the Red List of Brazilian Flora. Objective: To characterize morpho-anatomical aspects regarding germination and post-seminal development from C. crispa seeds; as well as studying the effect of cryopreservation on these seeds. Methods: We used light microscopy and electron microscopy to describe the microstructure of a 100 ripe seeds. We evaluated seed viability, seed germination, survival rate and protocorm weight in cryopreserved and non-cryopreserved material, with four replicas per treatment using 20 mg of plant material. Results: The seeds are fusiform, whitish yellow with a length from 700 to 900 µm and a water content of 5 %. Germination began seven days after sowing, the formation of the globular protocorm at 30 days and the formation of the seedling occurred 150 days. The persistent seed coat can compress the protocorm and cause it to collapse. The cryopreserved seeds presented 87.15 % viability, 78.32 % germination, 8.48 % survival and protocorms with 104.27 mg five months after sowing. Data wasn't different to non-cryopreserved seeds. Conclusions: The cryocapability of the seeds shows that cryopreservation can be used for long-term conservation. The results of this work contribute to the overall biology of C. crispa and to the propagation and storage of genetic material for conservation purposes.

Introducción: Cattleya crispa es una orquídea epífita ornamental con distribución geográfica restringida a la Mata Atlántica brasileña. Debido al extractivismo depredador y a la pérdida de hábitat inducida por el hombre, esta especie aparece en la Lista Roja de la Flora Brasileña. Objetivo: Caracterizar aspectos morfoanatómicos de la germinación y desarrollo inicial de semillas de C. crispa; así como estudiar el efecto de la criopreservación de estas semillas. Métodos: Utilizamos microscopía óptica, microscopía electrónica de barrido y microscopía electrónica de transmisión para describir la microestructura en 100 semillas maduras. Evaluamos la viabilidad de la semilla, la germinación de la semilla, la tasa de supervivencia y el peso de los protocormos en el material criopreservado y no criopreservado, con cuatro réplicas por tratamiento de 20 mg de material vegetal. Resultados: Las semillas son fusiformes, amarillo blanquecinas, con una longitud de 700 a 900 µm y un contenido de agua del 5 %. La germinación comenzó siete días después de la siembra, la formación del protocormo globular a los 30 días y la formación de la plántula a los 150 días. La cubierta de semilla persistente puede comprimir el protocormo y provocar su colapso. Las semillas criopreservadas presentaron 87.15 % de viabilidad, 78.32 % de germinación, 8.48 % de supervivencia y protocormos con 104.27 mg a los cinco meses de la siembra. Los datos no fueron diferentes a las semillas no criopreservadas. Conclusiones: La capacidad criogénica de las semillas muestra que la crioconservación puede utilizarse para la conservación a largo plazo. Los resultados de este trabajo contribuyen a la biología general de C. crispa y a la propagación y almacenamiento de material genético con fines de conservación.

Efficient propagation with in vitro seed germination of Vanda falcata / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Vanda falcata (Thunb.) Beer (Orchidaceae), a famous native orchid of China, Japan, and Korea, is known as one of the most beautiful and charming orchid species in the world (Ohwi, 1965; Lawler, 1984; Arditti, 2008). V. falcata is widely cultivated and delights the world with its compact plant shape, elegant white blooms, and sweet coconut-like scent. However, vegetative propagation by division has limited the development of V. falcata because of its inefficiency (Mitsukuri et al., 2009a, 2009b).

Agrobacterium-mediated transformation of the wild orchid Cattieya maxima Lindi / Transformación mediada por Agrobacterium de la orquídea silvestre Cattleya maxima Lindl / Transformação da orquídea silvestre Cattleya maxima Lindl mediada por Agrobacterium

Abstract Protocorms are unique anatomical structures; they are akin to rhizoids and are formed by young orchid seedlings under physiological conditions. Explanted orchid tissues produce similar structures called protocorm-like bodies (PLBs) when exposed to appropriate in vitro growing conditions. Both the propagative nature of PLBs and the easiness by which they can be generated, make these structures an attractive alternative to seed-mediated production for growing large numbers of plants. To increase somatic embryogenesis and optimize the procedure, PLBs of Cattleya maxima were transformed using the Agrobacterium tumefaciens method. The T-DNA carried a Hygromycin-resistance gene, a visible marker (GFP5-GUSA) and a rice gene encoding the Somatic Embryogenesis Receptor Kinase, deemed to be important for somatic embryogenesis. Treated PLBs generated somatic embryos developing Hygromycin-resistant plantlets. The insertion of T-DNA was confirmed by PCR, and GFP expression was observed using a fluorescent stereomicroscope. Transformed Cattleya maxima PLBs were more efficient in forming somatic embryos (60 - 80%) than untransformed controls (45 - 57%), and this contrast was maximized in hormone-free, Murashige and Skoog (MS) medium (80% of the transformed plants compared to 57% of the untransformed ones). This finding supports the notion that SERK plays an important role in Orchid embryogenesis.

Resumen Los protocormos son estructuras anatómicas únicas: son similares a los rizoides y se forman por vástagos jóvenes de orquídeas bajo condiciones fisiológicas. Los tejidos explantados de orquídeas producen estructuras llamadas Cuerpos Similares a Protocormos (PLBs) cuando están expuestos a condiciones apropiadas de crecimiento in vitro. Tanto la naturaleza propagativa de los PLBs como la facilidad con que se generan, hacen de estas estructuras una alternativa atractiva, frente a la mediada por semillas, para la producción de gran número de plantas en crecimiento. Para aumentar la embriogénesis somática y optimizar el procedimiento, se transformaron PLBs de Cattleya maxima usando el método de Agrobacterium tumefaciens. El T-DNA portaba un gen de resistencia a la Higromicina, un marcador visible (GFP5-GUSA) y un gen de arroz que codificaba para el receptor tipo quinasa de embriogénesis somática (SERK), considerado importante en la embriogénesis somática. Los PLBs tratados generaron embriones somáticos y desarrollaron plántulas resistentes a la Higromicina. La inserción del T-DNA se confirmó por PCR, y la expresión de GFP se observó usando un estereomicroscopio fluorescente. Los PLBs transformados de Cattleya maxima fueron más eficientes en desarrollar embriones somáticos (60-80%) que los controles no transformados (45-57%) y este contraste se maximizó en medio Murashige y Skoog (MS) libre de hormonas (80% de las plantas transformadas en comparación con 57% de las no transformadas). Estos hallazgos apoyan la noción de que SERK juega un papel importante en la embriogénesis de orquídeas.

Resumo Os protocormos são estruturas anatômicas únicas: são similares aos rizoides e se formam por hastes jovens de orquídeas sob condições fisiológicas. Os tecidos explantados de orquídeas produzem estruturas chamadas Corpos Similares a Protocormos (PLBs) quando estão expostos a condições apropriadas de crescimento in vitro. Tanto a natureza propagativa dos PLBs como a facilidade com que se generam, fazem com que estas estruturas sejam uma alternativa atrativa, comparativamente a mediada por sementes, para a produção de grandes números de plantas em crescimento. Para aumentar a embriogênesis somática e otimizar o procedimento, se transformaram PLBs de Cattleya maxima utilizando o método de Agrobacterium tumefaciens. O T-DNA carregava um gen de resistencia a Higromicina, um marcador visível (GFP5-GUSA) e um gen de arroz que codificava para o receptor tipo quinasa de embriogênesis somática (SERK), considerado importante na embriogênesis somática. Os PLBs tratados geraram embriões somáticos e desenvolveram plântulas resistentes a Higromicina. A inserção do T-DNA se confirmou por PCR, e a expressão de GFP se observou utilizando um estereomicroscópio de fluorescência. Os PLBs transformados de Cattleya maxima foram mais eficientes em desenvolver embriões somáticos (60-80%) que os controles não transformados (45-57%) e este contraste se potencializou em meio Murashige y Skoog (MS) livre de hormônios (80% das plantas transformadas em comparação com 57% das não-transformadas). Estes resultados apoiam a noção de que SERK desempenha um papel importante na embriogênesis de orquídeas.

Protocorm proliferation and regeneration of Dendrobium officinale / 中草药

Objective To establish an efficient tissue culture and rapid propagation system, and study the protocorm proliferation and regeneration conditions using seeds as explants in Dendrobium officinale. Methods Seeds of D. officinale were used as explants, protocorm was induced on inducement medium. After proliferation, the protocorm were transferred to the regeneration medium. Then the regenerated shoots were transferred into rooting medium to induce rooting of plantlets, and developing complete plant. Results The basic culture medium for D. officinale growth was 1/2 MS; The best culture medium formula for inducing protocorms was 1/2 MS +1.0 mg/L 6-BA + 0.2 mg/L NAA + 50 g/L mashed potato; The optimal proliferation medium for protocorm was 1/2 MS + 1.0 mg/L 6-BA + 0.5 mg/L NAA, and the maximum multiplication factor could reach 23. The optimal regeneration medium was 1/2 MS + 2.0 mg/L 6-BA + 0.2 mg/L NAA, regeneration rate can reach 95%; The best culture medium for seedling rooting was 1/2 MS + 0.3 mg/L NAA + 50 g/L potato juice, and rooting rate reached 100%. Conclusion This research provides an effective way for keeping good varieties of features and rapid propagation of D. officinale, at the same time helps to solve some theoretical problems in factory production of D. officinale.

Morphological and cytohistological observations of seed germination and protocorm development of Bletilla striata / 中国中药杂志

In order to investigate the mechanism of growth for Bletilla striata, which could be applied for rapid propagation, morphological and cytohistological of seed germination and protocorm development in vitro culture were observed using paraffin section techniques. In this study, we have found that the development of B. striata goes through four stages： embryo, protocorm, rhizome and pseudobulb. The end away from embryo suspensor is able to differentiate green buds after the seed of B. striata swelling, growing point. At the same time, the other end of embryo grows many white villous roots, with the green bud differentiating into cotyledon, the embryo breaking through seed coat and being protocorm. The shoot apical meristem of protocorm consists of tunica, corpus and leaf primordium, whose developmental flowing tunica-corpus theory. After more vascular bundle appeared from the leaf primordium, B. striata grows into the stage of rhizome. While in the stage of rhizome, the root primordium of tissue culture seedlings are differentia initially that derived from rhizome vascular bundle, belonging to internal origin. Subsequently, the pseudobulb forms by the inner meristem growing into mature parenchymatous tissue and the rhizome enlargement gradually.

Micropropagation of Brasilidium forbesii (Orchidaceae) through transverse and longitudinal thin cell layer culture / Micropropagação de Brasilidium forbesii (Orchidaceae) pela técnica 'thin cell layer' longitudinal e transversal

An efficient in vitro propagation method was established for Brasilidium forbesii (Hook.) Campacci using transverse and longitudinal thin cell layer (tTCL and lTCL, respectively) culture systems. Six-month-old protocorms from in vitro germinated seeds were used for this study. TCLs (1.0-mm thick) from protocorms were grown on Woody Plant Medium (WPM) supplemented with benzyladenine (BA) (0.5­4.0 µM).The lTCL technique was more efficient for inducing protocorm-like bodies (PLBs) and regenerating shoots than the tTCL technique. The frequency of PLB formation was influenced by BA concentration, and the lTCL explant grown on a medium containing 2.0 µM BA produced the highest percentage of new protocorms (77%) with a total of 22.7 PLBs per explant, after the first subculture on the same medium. Plantlet development was optimal on WPM medium containing 3.0 g L -1 activated charcoal, and indole-3 -butyric acid was not necessary for rooting. Regenerated plants were successfully acclimatized in a greenhouse after 16 weeks using vermiculite as the substrate (100% survival).

Foi estabelecido um método eficiente de propagação in vitro para Brasilidium forbesii (Hook.) Campacci utilizando a técnica 'thin cell layer' transversal (TCLt) e longitudinal (TCLl). Foram utilizados protocormos de seis meses obtidos da germinação in vitro. TCLs (1,0 mm de espessura) dos protocormos foram cultivados no meio 'Woody Plant Medium' (WPM), acrescido com benziladenina (BA) (0,5 a 4,0 µM). A técnica TCLl foi mais eficiente para indução de estruturas semelhantes a protocormos (ESPs) e regeneração de brotações do que a técnica TCLt. A frequência de formação de ESP foi influenciada pela concentração de BA e o explante TCLl, cultivado em um meio contendo 2,0 µM BA, produziu a mais alta percentagem de novos protocormos (77%), com um total de 22,7 PLBs por explante, após o primeiro subcultivo para o mesmo meio. O desenvolvimento das plântulas foi eficiente no meio WPM contendo 3,0 g L -1 de carvão ativado e o ácido indol-3- butírico (AIB) não foi necessário para o enraizamento. Plantas regeneradas foram estabelecidas com sucesso em casa de vegetação, utilizando vermiculita como substrato (100% de sobrevivência), após 16 semanas.

Study on Influencing Factors for Proliferation and Differentiation of Dendrobium officinale Protocorm-like Body / 广州中医药大学学报

Objective To screen the optimal medium formula for obtaining high proliferation rate and differentiation rate of protocorm-like body ( PLB) of Dendrobium officinale Kimura et Migo ( D.officinale) . Methods We observed the effect of different concentrations of phytohormones such as 6-benzylaminopurine (6-BA) , 1-naphthaleneacetic acid ( NAA) , 2,4-dichlorophenoxyacetic acid ( 2,4-D) and kinetin ( KT) , or their combinations together with organic additives such as potato extractive ( PE) , banana extractive ( BE) , apple extractive ( AE) and coconut milk ( CM) on the proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo. Results 6-BA, NAA and KT at certain concentrations were beneficial to the proliferation and growth of PLB, and when the concentration of KT was 1.0 mg/L, 50-d PLB proliferation times reached to 7.20. 2,4-D had obvious inhibitory effect on PLB proliferation and growth. The combination of 6-BA, NAA and KT showed stronger effect on PLB proliferation than the phytohormones used alone, and the combination of 0.5 mg/L 6-BA, 1.0 mg/L KT and 1.0 mg/L NAA had the best effect on PLB proliferation, 50-d PLB proliferation times reaching to 9.52. Certain concentrations of PE and AE could promote PLB differentiation, and 100 g/L PE brought well-grown seedling in test tube and the highest differentiation rate, being up to 92.6% . Conclusion The optimal hormone matching and additives concentration on proliferation and differentiation of PLB of Dendrobium officinale Kimura et Migo have been obtained, which can lay a foundation for its rapid propagation and artificial seed preparation.

In vitro regeneration and ploidy level analysis of Eulophia ochreata Lindl.

Various parameters including explant-type, medium compositions, use of phytohormones and additives were optimized for direct and indirect regeneration of E. ochreata, a medicinal orchid under threat. Protocorm-like-bodies (PLBs) proved to be the best explants for shoot initiation, proliferation and callus induction. Murashige and Skoog’s (MS) medium containing 2.5 mg L-1 6-benzylaminopurine (BAP), 1.0 mg L-1 kinetin (Kin) and additives (adenine sulfate, arginine, citric acid, 30 mg L-1 each and 50 mg L-1 ascorbic acid) was optimal for shoot multiplication (12.1 shoots and 7.1 PLBs per explant with synchronized growth), which also produced callus. Shoot number was further increased with three successive subcultures on same media and ~40 shoots per explant were achieved after 3 cycles of 30 days each. Additives and casein hydrolysate (CH) showed advantageous effects on indirect shoot regeneration via protocorm-derived callus. Optimum indirect regeneration was achieved on MS containing additives, 500 mg L-1 CH, 2.5 mg L-1 BAP and 1.0 mg L-1 Kin with 30 PLBs and 6 shoots per callus mass (~5 mm size). The shoots were rooted (70% frequency) on one by fourth-MS medium containing 2.0 mg L-1 indole-3-butyric acid, 200 mg L-1 activated charcoal and additives. The rooted plantlets were hardened and transferred to greenhouse with 63% survival rate. Flow-cytometry based DNA content analysis revealed that the ploidy levels were maintained in in vitro regenerated plants. This is the first report for in vitro plant regeneration in E. ochreata.

Mass propagation of Cymbidium giganteum Wall. ex Lindl. using in vitro seedlings.

In vitro seedlings were used as explants for protocorm like bodies (PLBs) production which in turn were used for regeneration purpose. PLBs were induced from the base of seedlings (1.0-1.5 cm in size) in MS + BAP (8.88 µM). After 90 days of inoculation, PLBs production rate started declining and most of the PLBs turned into plantlets. Preculture of seedlings in 1.0 µM thidiazuron (TDZ) for 7 days and transfer to BAP supplemented medium resulted in production of 16 PLBs per seedling within 90 days of culture. Increase of TDZ concentration to 2.5 µM and preculture time 15 days, resulted in induction of highest number of PLBs (19 PLBs per seedling) in the basal medium. The results emphasized the importance of thidiazuron (TDZ) concentration and preculture time for PLBs proliferation from the base of seedlings. The PLBs thus produced were used for regeneration studies. Irrespective of single, segmented or clumps of PLBs, the regeneration response was 100% in 2,4-D (4.52 µM) and KN (4.64 µM) but when KN was replaced by BAP (8.88 µM), response was observed only in clumps of PLBs, whereas in single and segmented ones it was 99 and 97%, respectively. Regenerants developed stout root system in half strength M medium supplemented with 2.84 µM of IAA and transferred to greenhouse with 90% survival. The present study holds tremendous potential as the mother plant is not destroyed and PLBs are produced as a continuous system.

In vitro regeneration of Coelogyne nervosa A.Rich. and Eria pseudoclavicaulis Blatt., threatened orchids of Western Ghats, India.

The seeds of C. nervosa and E. pseudoclavicaulis were germinated asymbiotically on Knudson C (KC) and Schenk and Hildebrandt basal medium (SH). Growth regulators such as 2,4-Dichlorophenoxyacetic acid (2,4-D) individually and in combinations with benzyladenine (BA) and kinetin were used for callus induction from the protocorm like bodies. Coelogyne nervosa showed maximum (90%) callus induction in Knudson C medium supplemented with 2,4-D (2.26 µM) and Eria pseudoclavicaulis showed 60% callus induction in Schenk and Hildebrandt medium supplemented with 2,4-D (2.26 µM). Calli developed a route of production of protocorm-like bodies and eventually developed into plantlets on transfer to growth regulator free half strength basal medium. The well rooted plants were hardened successfully in the potting mixture containing coconut husk, charcoal, and brick pieces in the ratio 2:1:1.

Effect of different culture conditions on protocorm-like bodies of Dendrobium candidum in bioreactor culture / 中草药

Objective: To explore the effect of different culture conditions on protocorm-like bodies (PLBs) of Dendrobium candidum in bioreactor culture. Methods: Inoculation quantity, ventilation, sucrose concentration, and light intensity were taken to carry out the single factor experiment design. Biomass was determined by drying method, polysaccharide content by phenol-sulfuric acid method, and SPSS16.0 software was used for data analysis. Results: Under the bioreactor culture, the nutrient medium components for the PLBs proliferation of D. candidum and polysaccharide accumulation were 1/2 MS medium supplemented with 1.0 mg/L NAA, 5% coconutmilk, and 3% sucrose, pH value was 6.0. The culture conditions, such as inoculation with 40 g/L, 15 μm pore porous nozzle, aeration volumes by 1.0 and 0.5 L/min used alternately, and photon flux of 2000 lx could improve the proliferation coefficient and polysaccharide contents of PLBs. Conclusion: Different culture conditions have the significant effect on the growth of PLBs and the accumulation of polysaccharide. Suitable culture conditions have the very important practic significance to the biomass of PLBs and main medicinal ingredient production of PLBs.

Research and application of Dendrobium officinale protocorm/protocorm-like bodies / 中国药学杂志

OBJECTIVE: To review the research progress of Dendrobium officinale protocorni/protocorm-like bodies according to relevant references at home and abroad in recent len years, and to explore its development prospect. METHODS: Based on the searched literatures, the research progress in the aspects of culture techniques, chemical constituents and pharmacological effects were summarized. RESULTS AND CONCLUSION: There had been a lot of researches on D. officinale protocorm/PLBs culture techniques, and D. officinale protocorm/PLBs could be used for resource regeneration and production of active polysaccharides, but its material basis and pharmacological activity still need to be further studied.

Study on chemical components of Dendrobium officinale protocorm / 中国药学杂志

OBJECTIVE: To study the chemical constituents of Dendrobium officinale protocorm. METHODS: The compounds were isolated and purified by means of column chromatography, and their structures were identified on the basis of spectroscopic data and comparison with standard literature data. RESULTS: Eight compounds were isolated from the(CH3)2CO, CHCl3 and 95% EtOH fractions of the EtOH extract, and identified as 1-O-p-feruloyl-β-D-glucopyranoside(I), arillatose B(II),4-β-D-glucopyranosy-loxy)benzyl alcohol(III), 4-hydroxymethyl-2,6-dimethoxyphenyl-β-D-glucopyranoside(IV), n-hexatriacontanoic acid(V), n-heptaco-sanol(VI), β-sitosterol(VII), and cycloart-23-ene-3β,25-diol(VIII). CONCLUSION: All these compounds were isolated from the protocorms of D. officinale for the first time. Compounds I, II, III, IV, and VIII were obtained from Dendrobium genus for the first time.

Seed germination and young seedling propagation of Anoectochilus roxburghii / 中国药学杂志

OBJECTIVE: To study the effect of different media on seed germination and young seedling propagation of Anoectochilus roxburghii. METHODS: A. roxburghii seeds were cultured in 6 basal media for 12 weeks, and germination rates were calculated; orthogonal design was used to study the effects of NAA and 6-BA on the propagation of young seedlings of A. roxburghii. After 4 months, the leaf number, stem internode number, root number, the length of root, plant height, and fresh mass were measured respectively. RESULTS: The effects of different media on seed germination and protocorm differentiation of A. roxburghii were different. Germination rate was the highest on MS medium, which was 60.38%, and protocorm differentiation was optimal on half-strength MS medium. Different combinations of NAA and 6-BA didn't show significant difference on the growth of young seedlings of A. roxburghii, and the appropriate medium was 1/2MS + NAA 1.0 g · L-1 + 6-BA 2.0 mg · L-1. CONCLUSION: The optimization of culture medium can stimulate seed germination and young seedling propagation of A. roxburghii. Copyright 2012 by the Chinese Pharmaceutical Association.

Propagación in vitro de Phalaenopsis (Orchidaceae) a partir de protocormos, mediante el sistema de inmersión temporal RITA / In vitro propagation of Phalaenopsis (Orchidaceae) from protocorms, using the temporary inmersion system RITA

Protocormos in vitro de Phalaenopsis de tres meses de edad se transfirieron a contenedores RITA® con el fin depropagarlos masivamente. Los factores evaluados fueron la concentración de sacarosa en el medio y la frecuencia de inmersión. Se dispusieron cinco pares de contenedores RITA® con medio de cultivo líquido a concentraciones de sacarosa de 0, 15, 30, 45 y 60 g/L. El medio utilizado fue el MS a la mitad de la concentración de las sales, suplementado con vitaminas y tidiazuron (5 mg/L). El experimento se realizó en dos etapas, cada una con duración de dos meses. La primera etapa con una frecuencia de inmersión de cuatro horas y la segunda con una frecuencia de inmersión de ocho horas, ambas con un tiempo de inmersión de un minuto. Los resultados mostraron que la mejor respuesta proliferativa, con 8,2 protocormos adventicios por protocormo por mes, se obtuvo en el medio con 15 g/L de sacarosa y un tiempo de inmersión de un minutocada cuatro horas.

In order to massively propagate Phalaenopsis orchids, three months old in vitro protocorms were transferred to RITA® vessels. The evaluation factors were the sucrose concentration in the culture medium and the frequency immersion. There was a set of five pairs of RITA® vessels with liquid culture medium containing sucrose at 0, 15, 30, 45, 60 g/L. A half-strength MS medium plus vitamins, supplemented with vitamins and thidiazuron (5 mg/L) was used. The experiment had two stages, each lasting two months. Each stage had a one minute immersion. The first stage had a four hour immersion frequency and the second one had an eight hour immersion frequency. Results showed that the best proliferating response was reached in a 15 g/L of sucrose medium with one minute of immersion time every four hours, resulting in 8,2 adventitious protocorms per protocorm per month.

Suspension culture of protocorm in Dendrobium candidum / 中草药

Object To study the feasibility of suspension culture of protocorm in Dendrobium candidum Wall ex Lindl and effect of inoculum and medium volume on the growth of protocorm Methods Effect of four basic media MS, 1/2 MS, 67 V, and B 5, inoculum and medium volume on the growth of protocorm were studied by completely random experimental design and orthogonal test design Results The growth of D candidum protocorms in liquid medium was markedly better than that in solid medium (P0 05), B 5 was much better than 1/2 MS (P

Tissue culture and clonal propagation of Gastrodia elata / 中草药

Object To develop a new method for the culturing of Gastrodia B1 in vitro, which may provide the theoretical basis of clonal propagation for its rapid reproduction Methods The small stem tubers and the stem buds were used as explants to culture in vitro under sterile conditions In the 1/2 MS medium containing 6 BA 1 mg/L, NAA 0 5 mg/L and banana 50 mg/L, the small stem tuber was induced to form protocorm In the 1/2 MS medium containing 6 BA 2 mg/L and NAA 0 2 mg/L, the stem bud was induced to form protocorm Results Each stem tuber formed a new protocorm within 50 d, which can be separated again to form rosette protocorm within 70 d The stem bud was cultured in vitro to form the protocorm within 140 d The protocorm bloomed within 160 d Conclusion The protocorm of G elata may be induced from the stem tuber and stem bud By subdividing these rosette protocorms a virtually indefinite clonal propagation can be achieved

Studies on cultivation of Gastrodia elata to lower expense and increase yield / 中草药

Object As stated in the subject. Methods By field cultivation.Results In sexual reproduction, (1) short tree branches can be used instead of the conventional tree trunks. Sow four layers of seed on to two timber layers. The peretration rate of Amillaria mellea (Vahl. ex Fr.) Karst into the germinated Gastrodia elata Bl.protocorm can attain well over 50%. After half year, the yield of both small and medium sized gastrodia tubers (2) Sow four layers of seed onto every two layers of long tree trunks may double the yield. In asexual propagation, the use of short tree branches not only can save timber material but also maintain the yield.Conclusion This new cultivation method can be used to save timber while maintaining the quality and quantity in the production of G. elata.

INFLUENCES OF FUNGUS MYCENAE SP.ON ACTIVITIES OF SOME ENZYMES AND EXTRACELLULAR PH OF PROTOCORMS OF DENDROBIUM CANDIDUM / 微生物学通报

An elicitor from fungus Mycenae sp. enhanced the extracellular pH of protocorms of Dendrobium candidum in two stages and also inspired the activities of PAL, POD and LOX. The different elicitors were different in enhancing the pH. The activities of PAL and POD ascended twice after elicitors were applied. The protocorms treated twice by elicitor had the higher PAL activity.

Antioxidant activity of crude and purified polysaccharide from suspension-cultured protocormns of Dendrobium candidum in vitro / 中成药

AIM: To study the antioxidant activity of crude polysaccharide(DCPP) and purified polysaccharide DCPP3c-1 from suspension-cultured protocorms of Dendrobium candidum in vitro. METHODS: The scavenging effect of crude and purified polysaccharide on radical was tested by 10-phenantthroline-Fe~(2+)-H_2O_2 systems and ammonium peroxydisulfate/ N,N,N′,N′-tetramethylethanediamine systems.The content of malonaldehyed(MDA) in mice liver homogenate and mice liver mitochondria were measured by TBA assay,and the swelling extent of mice liver mitochondria was also detected by spectrophotometric methods. RESULTS: DCPP and DCPP3c-1 could significantly scavenge ?OH and O~-_2,and the polysaccharide on ?OH showed dose-effect relationship.The 50% inhibitory concentrations(IC_(50)) of DCPP and DCPP3c-1 scavenging ?OH were 0.982 mg/mL and 0.930 mg/mL,and the IC_(50) of DCPP scavenging O~-_2 was 0.619 mg/mL.DCPP3c-1-1 had the highest inhibitory rate on O~-_2 at the concentration of 0.615 mg/mL,and then the inhibitory rate decreased with the increasing of its concentration.DCPP and DCPP3c-1 had inhibitory effects on lipid peroxidation in mice liver homogenate produced by itself and induced by Fe~(2+) or H_2O_2,and especially DCPP had stronger activity than DCPP3c-1.DCPP and DCPP3c-1 could inhibit the production of MDA in mice liver mitochondria,and could also decrease the swelling of mice liver mitochondria induced by ?OH. CONCLUSION: Both DCPP and DCPP3c-1 have antioxidant effects.