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1.
Bioreactors provide a rapid and efficient plant propagation system for many agricultural and forestry species, utilizing liquid media to avoid intensive manual handling. Large-scale liquid cultures have been used for micropropagation through organogenesis or somatic embryogenesis pathways. Various types of bioreactors with gas-sparged mixing are suitable for the production of clusters of buds, meristems or protocorms. A simple glass bubble-column bioreactor for the proliferation of ornamental and vegetable crop species resulted in biomass increase of 3 to 6-fold in 3–4 weeks. An internal loop bioreactor was used for asparagus, celery and cucumber embryogenic cultures. However, as the biomass increased, the mixing and circulation were not optimal and growth was reduced. A disposable pre-sterilized plastic bioreactor (2–5-l volume) was used for the proliferation of meristematic clusters of several ornamental, vegetable and woody plant species. The plastic bioreactor induced minimal shearing and foaming, resulting in an increase in biomass as compared to the glass bubble-column bioreactor. A major issue related to the use of liquid media in bioreactors is hyperhydricity, that is, morphogenic malformation. Liquid cultures impose stress signals that are expressed in developmental aberrations. Submerged tissues exhibit oxidative stress, with elevated concentrations of reactive oxygen species associated with a change in antioxidant enzyme activity. These changes affect the anatomy and physiology of the plants and their survival. Malformation was controlled by adding growth retardants to decrease rapid proliferation. Growth retardants ancymidol or paclobutrazol reduced water uptake during cell proliferation, decreased vacuolation and intercellular spaces, shortened the stems and inhibited leaf expansion, inducing the formation of clusters. Using a two-stage bioreactor process, the medium was changed in the second stage to a medium lacking growth retardants to induce development of the meristematic clusters into buds or somatic embryos. Cluster biomass increased 10–15-fold during a period of 25–30 days depending on the species. Potato bud clusters cultured in 1.5 1 of medium in a 2-l capacity bioreactor, increased during 10–30 days. Poplar in vitro roots regenerated buds in the presence of thidiazuron (TDZ); the biomass increased 12-fold in 30 days. Bioreactor-regenerated clusters were separated with a manual cutter, producing small propagule units that formed shoots and initiated roots. Clusters of buds or meristematic nodules with reduced shoots, as well as arrested leaf growth, had less distortion and were optimal for automated cutting and dispensing. In tuber-, bulb- and corm-producing plants, growth retardants and elevated sucrose concentrations in the media were found to enhance storage organ formation, providing a better propagule for transplanting or storage. Bioreactor-cultures have several advantages compared with agar-based cultures, with a better control of the contact of the plant tissue with the culture medium, and optimal nutrient and growth regulator supply, as well as aeration and medium circulation, the filtration of the medium and the scaling-up of the cultures. Micropropagation in bioreactors for optimal plant production will depend on a better understanding of plant responses to signals from the microenvironment and on specific culture manipulation to control the morphogenesis of plants in liquid cultures. 相似文献
2.
Theoretically, complete rejuvenation of mature trees should occur through somatic embryogenesis, however, this has not been
extensively studied. The main objective of the present study was to increase the efficiency of in vitro clonal propagation
for mature Quercus robur (100–300 years old), by induction of somatic embryogenesis as rejuvenation step prior to establishment of shoot culture through
micropropagation of somatic embryo-derived plantlets. Shoot culture lines of “mature” origin were established from epicormic
shoots of two centenarian oak genotypes (Sainza and CR-0) and maintained by axillary shoot proliferation. Embryogenic lines
were also initiated from epicormic leaf explants of the same genotypes and maintained by secondary somatic embryogenesis.
Although the frequency of somatic embryo conversion into plantlets was low in pedunculate oak, shoot culture lines could be
established and maintained by axillary branching from several germinated somatic embryos. For each genotype and shoot culture
line of the two origins (mature tree and somatic plantlets), shoot multiplication rate and elongation as well as rooting ability
parameters were compared. Compared with “mature-origin” shoot cultures and after more than one year propagation in vitro,
shoot lines established from somatic plantlets produced a significantly higher proportion of elongated, rootable shoots (from
26.0–31.6 to 36.8–40.5%) with increased rooting ability (from 3.3–45.6% to 23.2–89.8%). In the case of 300-year-old Sainza
genotype such a high organogenic capacity was similar to shoot cultures initiated from basal sprouts. Basal sprouts are considered
as “mature” material that retains juvenile characteristics compared with epicormic shoots forced from crown branches. Somatic
embryogenesis only slightly improved plant regeneration of shoot cultures from basal sprouts, thus validating their use as
“juvenile control”. The present results provide evidence that some rejuvenation occurred during the process of somatic embryogenesis
and resulted in improved shoot growth and rooting of somatic embryo-derived culture compared with “mature” shoot culture.
The results reported in this study might be useful in embryogenic systems with low plant conversion rates. The proposed experimental
model might also be useful in finding molecular markers of plant ontogeny. 相似文献
3.
Kee-Yoeup Paek Eun-Joo Hahn Sung-Ho Son 《In vitro cellular & developmental biology. Plant》2001,37(2):149-157
Summary The application of bioreactor culture techniques for plant micropropagation is regarded as one of the ways to reduce production
cost by scaling-up and automation. Recent experiments are restricted to a small number of species that, however, demonstrate
the feasibility of this technology. Periodic immersion liquid culture using ebb and flood system and column-type bubble bioreactors
equipped with a raft support system to maintain plant tissues at the air and liquid interface were found to be suitable for
micropropagation of plants via the organogenic pathway. Balloon-type bubble bioreactors proved to be fit for micropropagation
via somatic embryogenesis with less shear stress on cultured cells. Several cultivars of Lilium were successfully propagated using a two-stage culture method in one bioreactor. A large number of small-scale segments were
cultured for 4 wk with periodic immersion liquid culture to induce multiple bulblets from each segment, then the bulblet induction
medium was changed into bulblet growth medium by employing a submerged liquid bioreactor system. This culture method resulted
in a nearly 10-fold increase in bulblet growth compared to conventional culture with solid medium. About 20 000 cuttings of
virus-free potato could be obtained from 120 singlenode explants in a 20-liter balloon-type bubble bioreactor after 8 wk of
culture. The percentage of ex vitro survival and root induction of the cuttings was more than 95%. Other successful results were obtained from the micropropagation
and transplant production of chrysanthemum, sweetpotato, Chinese foxglove. Propagation systems via somatic embryogenesis in
Acanthopanax koreanum and thornless Aralia elata were established using a liquid suspension of embryogenic determined cells. More than 500 000 somatic embryos in different
stages were harvested from a 10-liter balloon-type bubble bioreactor after a 6-wk culture. Further development of these embryos
in solid medium and eventually in the field was successful. The bioreactor system could reduce initial and operational cost
for micropropagation, but further development of sophisticated technology might be needed to apply this system to plant micropropagation
industries. 相似文献
4.
Terrier B Courtois D Hénault N Cuvier A Bastin M Aknin A Dubreuil J Pétiard V 《Biotechnology and bioengineering》2007,96(5):914-923
The present article describes two novel flexible plastic-based disposable bioreactors. The first one, the WU bioreactor, is based on the principle of a wave and undertow mechanism that provides agitation while offering convenient mixing and aeration to the plant cell culture contained within the bioreactor. The second one is a high aspect ratio bubble column bioreactor, where agitation and aeration are achieved through the intermittent generation of large diameter bubbles, "Taylor-like" or "slug bubbles" (SB bioreactor). It allows an easy volume increase from a few liters to larger volumes up to several hundred liters with the use of multiple units. The cultivation of tobacco and soya cells producing isoflavones is described up to 70 and 100 L working volume for the SB bioreactor and WU bioreactor, respectively. The bioreactors being disposable and pre-sterilized before use, cleaning, sterilization, and maintenance operations are strongly reduced or eliminated. Both bioreactors represent efficient and low cost cell culture systems, applicable to various cell cultures at small and medium scale, complementary to traditional stainless-steel bioreactors. 相似文献
5.
Driven by the demands of the market and the manufacturing industry, disposable bioreactors have gained in importance in cell culture‐based processes during the last 10 years. Today they are widely accepted in R&D and also in manufacturing where process simplicity, safety and flexibility have top priority. Although disposable bioreactors are mainly used for cell expansions, glycoprotein secretions and virus generations realised with mammalian and insect cell lines, there are several reports delineating their suitability for the cultivation of plant cell and tissue cultures. This review describes the current disposable bioreactor types suitable for growing plant cell suspensions and organ cultures (hairy roots, meristematic clusters, somatic embryos) at Litre‐scale. Based on a definition of the term “disposable bioreactor”, a categorisation of the prevalent types for plant liquid cultures is presented. We describe the bioreactor regimes, working principles and bioengineering parameters of mechanically and pneumatically agitated bag bioreactors, which have advantages of process scalability and efficiency. Furthermore, results from the literature and data from our own research (obtained during production of undifferentiated bioactive cells, expressions of secondary metabolites and glycoproteins, and micropropagations of plant tissues) are discussed. 相似文献
6.
At Weyerhaeuser Company, somatic embryo production in liquid medium and manufactured seed delivery has been developed to reduce labor costs and increase efficiency of mass clonal propagation. We have scaled-up embryonal suspensor masses (ESM) of Douglas-fir in 1-l liquid medium flasks. Over 10,000 somatic embryos have been produced from a single flask. We have cryostored ESM of over 700 genotypes in liquid nitrogen. Somatic embryos of Douglas-fir have also been produced from liquid medium in a bioreactor. Different types of bioreactors are required for embryo multiplication and for cotyledonary embryo development. Over 250,000 Douglas-fir somatic seedlings from a large number of genotypes have been produced for clonal field tests. 相似文献
7.
Plant biotechnology can be used to conserve the germplasm of natural forests, and to increase the productivity and sustainability of plantations. Both goals imply working with mature trees, which are often recalcitrant to micropropagation. Conventional in vitro culture uses closed containers and gelled medium with sugar supplementation. Bioreactor culture uses liquid medium and usually incorporates aeration. The increased absorption of nutrients via the liquid medium together with the renewal of the air inside the bioreactors may improve the physiological state of the explants. In this review, we will explore the feasibility of using bioreactors to overcome the recalcitrance of many trees to micropropagation and/or to decrease the cost of large‐scale propagation. We will focus on the recent use of bioreactors during the multiplication, rooting (plant conversion in the case of somatic embryos), and acclimation stages of the micropropagation of axillary shoots and somatic embryos of forest trees (including some shrubs of commercial interest), in both temporary and continuous immersion systems. We will discuss the advantages and the main obstacles limiting the widespread implementation of bioreactor systems in woody plant culture, considering published scientific reports and contributions from the business sector. 相似文献
8.
Plant cell suspension cultures and hairy roots are potential sources of secondary metabolites and recombinant proteins. In
contrast to traditionally grown “whole wild plants” or “whole transgenic plants”, their production in bioreactors guarantees
defined controlled process conditions and therefore minimizes or even prevents variations in product yield and quality, which
simplifies process validation and product registration. Moreover, bioreactors and their configuration significantly affect
cultivation results by accomplishing and controlling the optimum environment for effective cell growth and production of bioactive
substances. This review highlights the main design criteria of the most widely used bioreactor types, both for plant cell
suspension cultures and for hairy roots, and outlines suitable low-cost disposable bioreactors which have found increasing
acceptance over the last 10 years.
Plants for human health in the post-genome era, PSE congress 26.8.2007–29.8.2007, Helsinki. 相似文献
9.
Regine Eibl Stephan Kaiser Renate Lombriser Dieter Eibl 《Applied microbiology and biotechnology》2010,86(1):41-49
Disposable bioreactors have increasingly been incorporated into preclinical, clinical, and production-scale biotechnological
facilities over the last few years. Driven by market needs, and, in particular, by the developers and manufacturers of drugs,
vaccines, and further biologicals, there has been a trend toward the use of disposable seed bioreactors as well as production
bioreactors. Numerous studies documenting their advantages in use have contributed to further new developments and have resulted
in the availability of a multitude of disposable bioreactor types which differ in power input, design, instrumentation, and
scale of the cultivation container. In this review, the term “disposable bioreactor” is defined, the benefits and constraints
of disposable bioreactors are discussed, and critical phases and milestones in the development of disposable bioreactors are
summarized. An overview of the disposable bioreactors that are currently commercially available is provided, and the domination
of wave-mixed, orbitally shaken, and, in particular, stirred disposable bioreactors in animal cell-derived productions at
cubic meter scale is reported. The growth of this type of reactor system is attributed to the recent availability of stirred
disposable benchtop systems such as the Mobius CellReady 3 L Bioreactor. Analysis of the data from computational fluid dynamic
simulation studies and first cultivation runs confirms that this novel bioreactor system is a viable alternative to traditional
cell culture bioreactors at benchtop scale. 相似文献
10.
Chin-Wen Ho Wei-Ting Jian Hui-Chun Lai 《In vitro cellular & developmental biology. Plant》2006,42(3):240-246
Summary
In vitro seedlings of Lilium × formolongi Hort. evs. Norikula, RaiZen No. 1, RaiZen No. 3, RaiZen Early, and Bailansa were used to induce callus by variously modified
Murashige and Skoog (MS) media, using protocols for flask culture and bioreactor culture. Green embryogenic callus proliferated
from roots near the base of bulblets of five varieties on media containing 0.53–5.3 μM α-naphthaleneacetic acid (NAA), and 28 cell lines were obtained by subcultures on the same medium. For flask culture, the
fresh weight (FW) of embryogenic cell clumps doubled every 4 wk on MS basal salts supplemented with 0.53°M NAA and 30 g l−1 sucrose. The maximum frequency of somatic embryos that developed into plantlets was 76.67±17% when plated onto solid MS basal
medium without plant growth regulators (PGRs). Among the treatments using four types of bioreactors, the best cell growth
and regeneration rate (74±0.14%) of somatic embryos was in a modified 2–1 bioreactor. Cells incubated in the other three bioreactors
furned brown and died. Histological study revealed that regeneration was by somatic embryogenesis. The regenerants showed
normal growth and flowering after 8–9 mo, in the field. A cell line of cv. Norikula has been subcultured in MS basal salts
containing 0.53 μM NAA every 2 mo. for 6 yr. The cell aggregates became more synchronous and many typical embryogenic cells with dense cytoplasm
were observed under a light microscope. The long-term embryogenic cells plated on MS basal medium still gave rise to numerous
somatic embryos and converted into plantlets. 相似文献
11.
In cassava, somatic embryogenesis starts with the culture of leaf explants on solid Murashige and Skoog-based medium supplemented
with auxins. Mature somatic embryos are formed within 6 wk. The cotyledons of the primary somatic embryos are used as explants
for a new cycle of somatic embryogenesis. The cotyledons undergo secondary somatic embryogenesis on both liquid and solid
Murashige and Skoog-based medium supplemented with auxins. Depending on the auxin, new somatic embryos are formed after 14–30
d after which they can be used for a new cycle of somatic embryogenesis. In liquid medium, more than 20 secondary somatic
embryos are formed per initial cultured embryo. In both primary and secondary somatic embryogenesis, the somatic embryos originate
directly from the explants. Transfer of clumps of somatic embryos to a Greshoff and Doy-based medium supplemented with auxins
results in indirect somatic embryogenesis. The direct form of somatic embryogenesis has a high potential for use in plant
propagation, whereas the indirect has a high potential for use in genetic modification of cassava.
Mature somatic embryos germinate into plants after desiccation and culture on a Murashige and Skoog-based medium supplemented
with benzylaminopurine (BA). Depending on the used BA concentration, plants can either be transferred either directly to the
greenhouse or after using standard multiplication protocols. 相似文献
12.
M. Capuana G. Petrini A. Di Marco R. Giannini 《In vitro cellular & developmental biology. Plant》2007,43(2):101-110
This is the first report on somatic embryogenesis in common ash (Fraxinus excelsior L.). Experiments on somatic embryogenesis induction were carried out on zygotic embryos at different phases of development
and maturation. The embryo axes were isolated and cultured on media containing different plant growth regulators (PGRs). Embryogenic
tissues were obtained from embryos collected at an incomplete maturation phase and cultured on a modified Murashige and Skoog
medium containing 8.8 μM 2,4-dichlorophenoxyacetic acid and 4.4 μM benzyl-adenine (BA). Embryos isolated from seeds at an
advanced stage of maturation showed only organogenetic phenomena. Embryogenic tissues were successfully subcultured and multiplied
on medium containing a reduced concentration of PGRs. After their isolation, somatic embryos were induced to develop and mature
by transfer to PGR-free medium and subsequent culture on medium containing 0.1 μM BA. Somatic embryos developed completely
and also germinated spontaneously. Embryo germination and conversion were significantly improved when subjected to a period
of storage at 4°C and transplant onto woody plant medium. Plantlets were successfully transferred to soil and acclimatized
in a “misted” greenhouse. 相似文献
13.
Diego Muñoz-Concha Sean Mayes Gracia Ribas Michael R. Davey 《Plant Cell, Tissue and Organ Culture》2012,109(1):123-130
The endangered Chilean tree Gomortega keule (Mol.) Baillon produces edible fruit, making it a potential crop. However, its cultivation from seed or cuttings is extremely
difficult. This paper reports the induction of somatic embryogenesis and the initiation of liquid cultures in this species.
Callus was induced from zygotic embryos and field-collected shoots. Somatic embryogenesis on zygotic embryos occurred at a
low frequency. Induction of somatic embryogenesis was accomplished on micropropagated shoots after 6.5 months on semi-solid
Murashige and Skoog (MS) medium with 30 g/l sucrose, 1.0 mg/l 2,4-dichlorophenoxyacetic acid and 1.0 mg/l 6-(γ,γ-dimethylallylamino)
purine (2iP). Liquid cultures of compact callus and small aggregates were obtained and showed optimum proliferation in MS
medium with 20 g/l sucrose, 0.01 mg/l α-naphthaleneacetic acid and 0.1 mg/l 2iP. The proliferation of friable embryogenic
callus was observed in liquid medium and will allow the propagation of selected genotypes of this tree on a large scale. Genetic
variation in two embryogenic genotypes cultured in vitro was not detected in an assessment using microsatellites; this approach
is suitable for tracing genotypes. 相似文献
14.
Hypocotyl segments ofEleutherococcus senticosuscultured on Murashigeand Skoog's (MS) medium with 4.5 µM2,4-D produced somaticembryos directly from the surface of explants without interveningcallus formation. When these somatic embryos were subculturedto the same MS medium with 4.5 µM2,4-D, friable embryogeniccalli were formed mainly from radicle tips of somatic embryos,but at a low frequency (5%). Selected embryogenic calli weremaintained on MS agar or liquid medium with 4.5 µM2,4-D.To induce somatic embryo development, embryogenic calli andcell clumps were transferred to MS medium lacking 2,4-D. Thefrequency of somatic embryo formation differed between culturetypes with 1570 embryos formed per Petri dish from callus cultureand 5514 embryos formed per flask from cell suspension cultures.Somatic embryos formed on agar medium had larger cotyledonsthan those of embryos formed in liquid medium. GA3treatmentwas necessary to induce germination from somatic embryos. Therate of plant conversion was 97% in somatic embryos from callusculture and 76% in embryos from liquid culture. Regeneratedplantlets were successfully acclimatized in the glasshouse.Copyright1999 Annals of Botany Company Eleutherococcus senticosus, micro propagation, somatic embryogenesis. 相似文献
15.
Somatic embryogenesis in soybean via somatic embryo cycling 总被引:4,自引:0,他引:4
Wennuan Liu Patricia J. Moore Glenn B. Collins 《In vitro cellular & developmental biology. Plant》1992,28(3):153-160
Summary The objectives of the present research were: a) to develop an efficient soybean embryogenic regeneration system characterized
by a high frequency of explant response and a large number of somatic embryos per explant; b) to evaluate the factors affecting
somatic embryogenesis via somatic embryo cycling; and c) to identify the origin of somatic embryos in the system. A highly
improved and efficient system for soybean somatic embryogenesis was established using somatic embryo cotyledons and somatic
embryo hypocotyl/radicle explants plated on α-naphthaleneacetic acid (NAA) or 2,4-dichlorophenoxyacetic acid (2,4-D) supplemented
MS basal media. The system included somatic embryo cycling between liquid and solid medium and it consistently gave rise to
a much higher frequency of explant response and a larger number of embryos per responding explant than those obtained from
zygotic cotyledon explant tissues. Genotype, differences were observed for response in some of the treatments with cv “Fayette”
being more responsive than “J103”. Histological studies revealed that somatic embryos induced in the somatic embryo cycling
system originated almost exclusively from epidermal cells on both 2,4-D and NAA inductive media. The cells of the epidermis
proliferated to produce somatic embryos directly without an intervening callus phase. A single-cell origin of somatic embryos
was observed in cultures on a 40 mg/liter 2,4-D treatment. A large number of responding cells in the epidermis was also observed
in the 10 mg/liter NAA treatment. The single-cell origin of somatic embryos from epidermal layers of the explant tissues should
facilitate development of an efficient transformation system for soybean. 相似文献
16.
Application of bioreactor systems for large scale production of horticultural and medicinal plants 总被引:1,自引:0,他引:1
Automation of micropropagation via organogenesis or somatic embryogenesis in a bioreactor has been advanced as a possible way of reducing costs. Micropropagation by conventional techniques is typically a labour-intensive means of clonal propagation. The paper describes lower cost and less labour-intensive clonal propagation through the use of modified air-lift, bubble column, bioreactors (a balloon-type bubble bioreactor), together with temporary immersion systems for the propagation of shoots, bud-clusters and somatic embryos. Propagation of Anoectochilus, apple, Chrysanthemum, garlic, ginseng, grape, Lilium, Phalaenopsis and potato is described. In this chapter, features of bioreactors and bioreactor process design specifically for automated mass propagation of several plant crops are described, and recent research aimed at maximizing automation of the bioreactor production process is highlighted. 相似文献
17.
Nazmul H. A. Mamun Cyrus K. Aidun Ulrika Egertsdotter 《In vitro cellular & developmental biology. Plant》2018,54(6):612-620
Somatic embryogenesis offers many benefits for clonal propagation in large-scale plant production of conifers. A key rate-limiting step is the conversion from early-stage somatic embryos in pro-embryogenic masses (PEMs) to the maturation stage. Immature embryos in PEMs are present at different developmental stages, where some are unable to respond to the maturation treatment, thus limiting yields of mature embryos. Synchronization of early somatic embryo development in PEMs could greatly improve subsequent yields of mature embryos. A temporary immersion bioreactor designed for Norway spruce (Picea abies (L.) H.Karst.) was used in this study. Through a specific system for dispersion, connected tissue of PEMs, composed of immature embryos grown in liquid medium in the temporary immersion bioreactors or on solid medium as a control, was dispersed and redistributed in a more uniform spatial arrangement. It was demonstrated that development of mature embryos could be significantly stimulated by dispersion, compared to controls, in both medium types. Synchronization of maturation was evaluated by a statistical approach. The present study shows that the yield of mature embryos from dispersed PEMs was three to five times higher than that from non-dispersed controls in three of four cell lines of Norway spruce tested, both in bioreactors and on solid medium. 相似文献
18.
Jing Li Yang Bo Zhao Eun Soo Seong Myong Jo Kim Won Hee Kang Na Young Kim Chang Yeon Yu Cheng Hao Li 《Plant biotechnology reports》2010,4(4):261-267
We describe culture conditions for a high-efficiency in vitro regeneration system of Papaver
nudicaule through somatic embryogenesis and secondary somatic embryogenesis. The embryogenic callus induction rate was highest when
petiole explants were cultured on Murashige and Skoog (MS) medium containing 1.0 mg l−1 α-naphthaleneacetic acid (NAA) and 0.1 mg l−1 6-benzyladenine (BA) (36.7%). When transferred to plant growth regulator (PGR)-free medium, 430 somatic embryos formed asynchronously
from 90 mg of embryogenic callus in each 100-ml flask. Early-stage somatic embryos were transferred to MS medium containing
1.0 mg l−1 BA and 1.0 mg l−1 NAA to germinate at high frequency (97.6%). One-third-strength MS medium with 1.0% sucrose and 1.0 mg l−1 GA3 had the highest frequency of plantlet conversion from somatic embryos (91.2%). Over 90% of regenerated plantlets were successfully
acclimated in the greenhouse. Secondary somatic embryos were frequently induced directly when the excised hypocotyls of the
primary somatic embryos were cultured on MS medium without PGRs. Sucrose concentration significantly affected the induction
of secondary embryos. The highest induction rate (89.5) and number of secondary somatic embryos per explant (9.3) were obtained
by 1% sucrose. Most secondary embryos (87.2–94.3%) developed into the cotyledonary stage on induction medium. All cotyledonary
secondary embryos were converted into plantlets both in liquid and on semisolid 1/3-strength MS medium with 1.0% sucrose. 相似文献
19.
The purpose of this study was to evaluate and compare the use of liquid and solid Murashige and Skoog (MS) medium in different culture vessels for mass production of Catharanthus roseus, an important source of anticancerous compounds, vincristine and vinblastine. Three media conditions i.e. agar-solidified medium (S), liquid medium in agitated conical flask (L) and growtek bioreactor (B) were used. Rapid propagation was achieved through in vitro somatic embryogenesis pathway. The process of embryogenesis has been categorized into induction, proliferation, maturation and germination stages. All in vitro embryogenesis stages were conducted by withdrawing spent liquid medium and by adding fresh MS medium. In optimized 4.52 μM 2,4-D added MS, the callus biomass growth was low in solid (1.65 g) compared to liquid medium in agitated conical flask (1.95 g) and in bioreactor (2.11 g). The number of normal somatic embryos was more in solid medium (99.75/50 mg of callus mass) compared to liquid medium used in conical flask (83.25/callus mass) and growtek bioreactor (84.88/callus mass). The in vitro raised embryos maturated in GA3 (2.60 μM) added medium; and in bioreactor the embryo growth was high, a maximum length of 9.82 mm was observed at the end of four weeks. These embryos germinated into seedlings in BAP (2.22 μM) added medium and the embryo germination ability was more (59.41%) in bioreactor compared to liquid medium in conical flask (55.5%). Shoot length (11.25 mm) was also high in bioreactor compared to agitated conical flask. The liquid medium used in agitated conical flask and bioreactor increased seedling production efficiency, at the same time it also reduced plant recovery time. The embryo generated plants grew normally in outdoor conditions. The exploitation of medium to large culture vessel or bioreactor may make the process more efficient in getting large number of Catharanthus plant as it is the only source of anti-cancerous alkaloids, vincristine and vinblastine.Abbreviations: BA, N6-benzyladenine; 2,4-D, 2,4-Dichlorophenoxyacetic acid; GA3, gibberellic acid; NAA, naphthalene acetic acid; MS, Murashige and Skoog (1962) medium; S, agar-solidified medium; L, liquid medium in agitated conical flask; B, growtek bioreactor 相似文献
20.
Large-scale growth and taxane production in cell cultures of Taxus cuspidata (Japanese yew) using a novel bioreactor 总被引:4,自引:0,他引:4
S. H. Son S. M. Choi Y. H. Lee K. B. Choi S. R. Yun J. K. Kim H. J. Park O. W. Kwon E. W. Noh J. H. Seon Y. G. Park 《Plant cell reports》2000,19(6):628-633
A novel type of bioreactor was successfully developed for the production of taxol and its precursors by culturing cells of
Taxus cuspidata (Japanese yew) on a pilot-scale. Rapidly growing cell lines were selected from callus cultures derived from immature embryos
of yew. The cells were inoculated in 20-l capacity bioreactors of different types to test the growth performance. The models
of small-scale bioreactors incorporated in this study included a balloon-type bubble bioreactor (BTBB), a bubble-column bioreactor
(BCB), a BCB with a split-plate internal loop, a BCB with a concentric draught-tube internal loop, a BCB with a fluidized
bed bioreactor, and two different models of stirred tank reactors. Among the reactors, BTBB appeared to be the most efficient
in promoting cell growth. The doubling time of cell growth in BTBB was 12 days with a 30% inoculation cell density. The optimum
time for medium replacement or feeding was 12–15 days after inoculation as determined by monitoring both the levels of sugars
and medium conductivity. When yew tree cells were grown in different sizes (100–500-l) of BTBBs, more than 70% cell viability
was recorded at the time of harvest. The growth pattern of the cells in the pilot-scale BTBB appeared to be the same as that
of cells in the 20-l bioreactors. Approximately 3 mg/l of taxol and 74 mg/l total taxanes were obtained after 27 days of culture.
Received: 6 April 1999 / Revision received: 23 August 1999 / Accepted: 31 August 1999 相似文献