Liquid Culture Systems for in vitro Plant Propagation

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522 E. Gontier et al. 3.6 Scale up of the system In order to scale up our temporary immersion system (TIS), we developed a 20-litre vessel composed of three Nalgene polypropylene flasks. This larger TIS allowed us to produce kilograms of biomass (Figure 12 F) with FCs levels comparable to those obtained in the 3-litre TIS (growth: 1.6 g DW day -1 and calculated FC productivity: 3.8 mg FCs day -1 l -1 ). This 20-litre size is the maximum that we could reach within the lab because of the size of our autoclave and of our laminar hood (in which the inoculation is performed). As compared to the results obtained in the 3-litre devices, heterogeneity in biomass aspect was similar (low chlorophyll content within the shoot core). Thus, we estimate that 20 l is the maximum vessel size (volume) in which we can cultivate Ruta shoots without the need for further light supply (i.e. development of a photobioreactor). 4. Conclusions Firstly, shoot cultures were initiated in liquid medium in 250 ml Erlenmeyer flasks. After studying growth and FCs production in these conditions, a 4-litre Setric TM bioreactor was inoculated with shoots and the culture parameter were studied. As expected, because of the presence of a metallic matrix in the tank, a part of the plant material was trapped in some parts of the reactor and a high heterogeneity of the biomass could be observed. Thus, we developed a home made bioreactor consisting in a simple glass jar coupled to a second one. The inoculation was performed by opening the screw cap under a laminar hood and the culture was performed in the major jar with air bubbled with a sparger (bubble flask). The coupling of the major jar to an annex one allowed us to transfer the culture medium from the first to the second one and to weigh this containing the biomass. This allowed us to measure growth in real time. This simple bioreactor also allowed us to carry out many experiments and repetitions in parallel and to show, for example, that a lower sucrose concentration (10 g l -1 instead of 30 g l -1 ) gave best results in terms of growth and FCs production. Furthermore, because temporary immersion has been described as a possible way to obtain a higher growth rate and less vitrification of the biomass, we built a very simple (and low cost) system to perform cultures in these conditions. The trials showed that temporary immersion lead to results equal or superior to those of permanent immersion regime. Shoot growth was improved and its quality (no vitrification, high FCs content) was better also. Then, based on our in-house technology (bubble flasks for permanent or temporary immersion regime) we evaluated the maximum biomass and FCs
Development and validation of low cost bioreactor 523 production that could be reached with such bioreactors. We showed that very high densities of Ruta shoots can be obtained and that a scale up step at 20 l can be performed successfully. This plant material contains large amounts of FCs that can be released when permeabilizing the biomass with detergent (Tween20). This process can be performed without killing the tissues, allowing further culture and permeabilization cycles. In conclusion, based on the experience we gained in the past on hairy root cultures (Boitel-Conti et al., 1995 a, b, 1996, 1997), we developed a home-made, simple and low costs bioreactor system that allowed a good biomass and FCs production. These results are of interest for potential applications of plant organ cultures for secondary metabolite production. They are also very promising for developing useful bioreactor systems for plant micropropagation (Gontier et al., 1993; Lorenzo et al., 1998, Hsiao et al., 1999). References Boitel-Conti M, Laberche JC, Gontier E & Sangwan-Norreel BS (1995a) Culture de racines transformées de Datura innoxia Mill. en bioréacteur: comparaison de 3 types de procédés. C.R. Acad. Sci. Paris 318: 593-598 Boitel-Conti M, Gontier E, Laberche JC, Ducrocq C and Sangwan-Norreel BS (1995b) Permeabilization of Datura innoxia hairy roots for release of stored tropane alkaloids. Planta Medica 61: 287-290 Boitel-Conti M, Gontier E, Laberche JC, Ducrocq C & Sangwan-Norreel BS (1996) Inducer effect of Tween 20 permeabilization treatment used for release of stored tropane alkaloids in Datura innoxia Mill. hairy root cultures. Plant Cell Reports 16: 241-244 Boitel M, Gontier E, Assaf C, Laberche JC, Ducrocq C & Sangwan-Norreel BS (1997) Procédés de production de métabolites primaires et secondaires à partir d'organes végétaux cultivés in vitro, Patent: France n°FR 97 00641 Bourgaud F, Gravot A, Milesi S & Guckert A (2001) Production of plant secondary metabolites: a historical perspective. Plant Science 161: 839-851 Dornenburg H & Knorr D (1995) Strategies for the improvement of secondary metabolite production in plant cell cultures. Enzyme and Microbial Technology 17: 674-684 Gamborg OL, Miller RA & Ojima V (1968) Nutrient requirements of suspension cultures of soybean root cells. Exp. Cell. Res. 50: 151-158 Gontier E, Clement A, Tran TLM, Lievre K, Gravot A, Guckert A & Bourgaud F (2002) Hydroponic combined with natural or forced root permeabilization: a promising technique for plant secondary metabolites production. Plant Science (in press) Gontier E, Fliniaux MA, Barbotin JN & Sangwan-Norreel BS (1993) Tropane alkaloid levels in the leaves of micropropagated Datura innoxia Mill. plants. Planta Medica 59: 432-435 Hsiao TY, Bacani FT, Carvalho EB & Curtis WR (1999) Development of a low capital investment reactor system: application for plant cell suspension culture. Biotechnol. Prog. 15: 114-122 Koppenhofer E (1995) Kaliumkanalblocker in der Neurologie: 5-MOP zur Behandlung MSbedingter Funktionsdefizite. TW Neurologie Psychiatrie: 585-591
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LIQUID CULTURE SYSTEMS FOR IN VITRO
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A C.I.P. Catalogue record for this
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Contents Contributing Authors xiii
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Liquid culture systems for in vitro
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Contributing Authors Adelberg, J. 4
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Preface Plant cell, tissue and orga
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Chapter 1 General introduction: a p
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General introduction 3 2. Cell susp
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General introduction 5 rooted in gr
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General introduction 7 (Winkelmann
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General introduction 9 the bioreact
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General introduction 11 (TIS), resu
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General introduction 13 Barz W, Rei
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General introduction 15 Hohe A, Win
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General introduction 17 Shimazu T &
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I. Bioreactors
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22 Wayne R. Curtis 1. Introduction
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24 Wayne R. Curtis headspace double
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26 Wayne R. Curtis that is required
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28 Wayne R. Curtis C L y � P O2
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30 Wayne R. Curtis since there is a
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32 Wayne R. Curtis 5 cm 30 cm 30 o
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34 Wayne R. Curtis Pˆ N � � me
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36 Wayne R. Curtis Radial Flow Impe
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38 Wayne R. Curtis CS = Medium diss
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40 Wayne R. Curtis Murashige T & Sk
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42 Anne Kathrine Hvoslef-Eide et al
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Chapter 4 Practical aspects of bior
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Practical aspects of bioreactor app
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Chapter 5 Simple bioreactors for ma
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Simple bioreactors for mass propaga
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96 K. Y. Paek et al. opportunity fo
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98 K. Y. Paek et al. disadvantages
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100 K. Y. Paek et al. 3.1 Dissolved
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102 K. Y. Paek et al. optimizing bi
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104 K. Y. Paek et al. cosmetics, wh
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106 K. Y. Paek et al. protocol, mor
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108 K. Y. Paek et al. use. In order
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110 K. Y. Paek et al. a b c d e f F
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112 K. Y. Paek et al. a b c d e f F
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114 K. Y. Paek et al. Escalona M, L
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116 K. Y. Paek et al. Son SH & Paek
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118 Seppo Sorvari et al. economical
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120 Seppo Sorvari et al. membrane w
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122 Seppo Sorvari et al. 3. Results
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124 Seppo Sorvari et al. D.O. 160 1
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Chapter 8 Cost-effective mass cloni
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Cost-effective mass cloning of plan
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144 Eun-Joo Hahn & Kee-Yoeup Paek 1
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146 Eun-Joo Hahn & Kee-Yoeup Paek F
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148 Eun-Joo Hahn & Kee-Yoeup Paek T
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150 Eun-Joo Hahn & Kee-Yoeup Paek 3
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152 Eun-Joo Hahn & Kee-Yoeup Paek E
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Chapter 10 Control of growth and di
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Control of growth and differentiati
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Chapter 11 Temporary immersion syst
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Temporary immersion system: a new c
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198 Elio Jiménez González 1. Intr
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200 Elio Jiménez González Several
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202 Elio Jiménez González Figure
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204 Elio Jiménez González exploit
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206 Elio Jiménez González weeks i
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208 Elio Jiménez González germina
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210 Elio Jiménez González Escalan
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Chapter 13 Use of growth retardants
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Use of growth retardants for banana
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Chapter 14 Somatic embryo germinati
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Somatic embryo germination of Psidi
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Somatic embryo germination of Psidi
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232 Tino Hempfling & Walter Preil N
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234 Tino Hempfling & Walter Preil 3
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236 Tino Hempfling & Walter Preil F
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238 Tino Hempfling & Walter Preil F
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240 Tino Hempfling & Walter Preil 4
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242 Tino Hempfling & Walter Preil R
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244 C. Damiano et al. Over the last
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246 C. Damiano et al. 2.2 Temporary
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248 C. Damiano et al. Table 3: Wate
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250 C. Damiano et al. hyperhydricit
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Chapter 17 Optimisation of growing
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Optimisation of growing conditions
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264 Katerina Grigoriadou et al. cul
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266 Katerina Grigoriadou et al. 2.4
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268 Katerina Grigoriadou et al. mic
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270 Katerina Grigoriadou et al. of
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272 Katerina Grigoriadou et al. 4.
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274 Katerina Grigoriadou et al. Tei
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276 Ch. Wawrosch et al. possesses s
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278 Ch. Wawrosch et al. 3. Results
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280 Ch. Wawrosch et al. References
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Chapter 20 Propagation of Norway sp
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Propagation of Norway Spruce 285 2.
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Propagation of Norway Spruce 287 su
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Propagation of Norway Spruce 289 5.
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Propagation of Norway Spruce 291 bl
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Propagation of Norway Spruce 293 H
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296 M. Vágner et al. 1995). In liq
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298 M. Vágner et al. maturation pe
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300 M. Vágner et al. mature somati
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302 M. Vágner et al. Acknowledgeme
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304 Christopher Hunter & Lionel Lev
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314 Michel Petitprez et al. 1. Intr
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316 Michel Petitprez et al. 2.2 QTL
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318 Michel Petitprez et al. Figure
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320 Michel Petitprez et al. Table 1
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322 Michel Petitprez et al. Gentzbi
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324 F. Afreen et al. 1. Introductio
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326 F. Afreen et al. precotyledonar
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328 F. Afreen et al. Dry mass (mg/e
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330 F. Afreen et al. under photoaut
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332 F. Afreen et al. Figure 3: a) C
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334 F. Afreen et al. the plant qual
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Chapter 25 Potential of flow cytome
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Potential of flow cytometry 339 dis
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Potential of flow cytometry 341 Fig
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Potential of flow cytometry 343 tha
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Chapter 26 Somatic embryogenesis of
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Somatic embryogenesis of Gentiana 3
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Chapter 27 Induction and growth of
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Induction and growth of tulip 361 a
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Induction and growth of tulip 363 T
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Chapter 28 Micropropagation of Ixia
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Micropropagation of Ixia 367 for PA
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Micropropagation of Ixia 369 Biomas
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Micropropagation of Ixia 371 Figure
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Chapter 29 Micropropagation of Rosa
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Micropropagation of Rosa 375 The BA
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Micropropagation of Rosa 377 Platfo
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Micropropagation of Rosa 379 3. Res
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Micropropagation of Rosa 381 The ro
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Micropropagation of Rosa 383 4. Dis
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Micropropagation of Rosa 385 Murash
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Chapter 30 Mass propagation of coni
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Mass propagation of conifer trees 3
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Chapter 31 Potentials for cost redu
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Potentials for cost reduction 405 c
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Potentials for cost reduction 407 c
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Potentials for cost reduction 409 s
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Potentials for cost reduction 411 N
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Potentials for cost reduction 413 G
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Chapter 32 A new approach for autom
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A new approach for automation 417 (
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A new approach for automation 419 p
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A new approach for automation 421 T
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A new approach for automation 423 D
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426 B. McAlister et al. 1. Introduc
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428 B. McAlister et al. 2.2 Multipl
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430 B. McAlister et al. Table 2: Mu
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432 B. McAlister et al. rest time -
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434 B. McAlister et al. Figure 3: M
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436 B. McAlister et al. Average mul
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438 B. McAlister et al. Table 4: Co
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440 B. McAlister et al. Semi-solid
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442 B. McAlister et al. Escalona M,
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444 Jeffrey Adelberg including Host
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446 Jeffrey Adelberg BioSystems (Ho
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448 Jeffrey Adelberg more prolific
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450 Jeffrey Adelberg vessel of liqu
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452 Jeffrey Adelberg Figure 4: Labo
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454 Jeffrey Adelberg constituted a
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456 Jeffrey Adelberg Acknowledgemen
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Chapter 35 Aeration stress in plant
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Aeration stress in plant tissue cul
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Page 472 and 473: 476 Hans R. Gislerød et al. 1. Int
Page 474 and 475: 478 Hans R. Gislerød et al. biorea
Page 476 and 477: 480 Hans R. Gislerød et al. A low
Page 478 and 479: 482 Hans R. Gislerød et al. Table
Page 480 and 481: 484 Hans R. Gislerød et al. common
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Page 488 and 489: 492 Hans R. Gislerød et al. Morten
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Page 502 and 503: V. Biomass for Secondary Metabolite
Page 504 and 505: 510 E. Gontier et al. 1. Introducti
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Page 518 and 519: 524 E. Gontier et al. Lorenzo JC, G
Page 520 and 521: 526 Dirk Wilken et al. 1. Introduct
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Page 524 and 525: 530 Dirk Wilken et al. described a
Page 526 and 527: 532 Dirk Wilken et al. packed cell
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Page 532 and 533: Chapter 40 Cultivation of root cult
Page 534 and 535: Cultivation of root cultures of Pan
Page 540 and 541: Chapter 41 Optimisation of Panax gi
Page 542 and 543: Optimisation of Panax ginseng liqui
Page 550 and 551: 558 S. M. Nalawade et al. performan
Page 552 and 553: 560 S. M. Nalawade et al. Plantlets
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Page 560 and 561: Production of taxanes 569 2. Materi
Page 562 and 563: Production of taxanes 571 Figure 2:
Page 564 and 565: Production of taxanes 573 various T
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Acknowledgments The editors thank t
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578 Contents 130, 131, 133, 134, 13
Page 570 and 571:
580 Contents poplar, see Populus Po
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582 Contents bubble aerated 165 bub
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584 Contents ginsenoside content, p
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586 Contents plant growth regulator
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588 Contents -free microcorms 71 -f
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