Design, Development, And Applications Of Mist Bioreactors For Micropropagation And Hairy Root Culture

Towler, Melissa J.; Kim, Yoojeong; Wyslouzil, Barbara E.; Correll, Melanie J.; Weathers, Pamela J.

doi:10.1007/978-1-4020-3694-1_7

Cited by 8 publications

(6 citation statements)

References 66 publications

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“…The optimization of cultivation depends on the type of bioreactor used and its operation, including aeration and immersion time, which is widely reported in the literature [ 60 , 61 ]. This has been demonstrated by us for both Schisandra chinensis [ 62 ] and Verbena officinalis cultures [ 63 ].…”

Section: Resultsmentioning

confidence: 99%

Antioxidant Potential and Enhancement of Bioactive Metabolite Production in In Vitro Cultures of Scutellaria lateriflora L. by Biotechnological Methods

et al. 2022

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Studies carried out using three different in vitro assays and a biological setting (Escherichia coil) demonstrated the antioxidant activity of Scutellaria lateriflora microshoot extract. Moreover, the extract exhibited no toxicity in a brine shrimp lethality bioassay. These results indicated that microshoots are a rich, safe source of antioxidants, which encouraged us to enhance their production in vitro. In agar and agitated cultures, two biotechnological strategies were applied: feeding the cultures with the biogenetic precursors of the phenolics—phenylalanine and tyrosine, and eliciting them with methyl jasmonate. Specific Scutellaria flavonoids and verbascoside were analysed by HPLC. Feeding with precursors (1 g/L) in agar cultures decreased the production of the metabolites. In agitated cultures, different concentrations of precursors (1.0–2.5 g/L) and the elicitor (10; 50; 100 µM) were tested. Additionally, parallel feeding with the precursor and elicitor in a concentration of 50 µM were applied. The best strategy for total flavonoid and verbascoside production was phenylalanine feeding (1.5 g/L), max. 3765 and 475 mg/100 g DW, respectively, after 7 days. This is the first report documenting the high antioxidant production in S. lateriflora microshoots after feeding with phenylalanine. Moreover, for the first time, bioreactor cultures were successfully maintained, obtaining attractive results (max. total flavonoid content 2348 and verbascoside 485 mg/100 g DW).

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Section: Resultsmentioning

confidence: 99%

Antioxidant Potential and Enhancement of Bioactive Metabolite Production in In Vitro Cultures of Scutellaria lateriflora L. by Biotechnological Methods

et al. 2022

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“…Earlier work with the mist reactor showed the benefits of growing hairy roots in mist compared to other reactors: healthier roots than in liquid systems (Towler et al, 2006(Towler et al, , 2007Wyslouzil et al, 2000), higher yields of secondary products (Kim et al, 2001), no oxygen limitations even in densely packed root beds (Weathers et al, 1999), and growth rates equal to or greater than in shake flasks (Towler et al, 2007). Serious limitations, however, existed: designs were prone to breakdown, all were not scalable, droplet delivery was tightly coupled to gas flow rate, and volumetric throughput was low (Table II).…”

Section: Discussionmentioning

confidence: 98%

“…Proteins offer additional challenges with the need to oxygenate roots in liquid adding shear forces that can degrade secreted proteins. Use of a gas-phase reactor like a mist reactor Towler et al, 2006;Weathers and Giles, 1988;Weathers and Wyslouzil, 2000) may help to alleviate external degradation of any secreted protein while still enhancing biomass yield.…”

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confidence: 99%

Production of mouse interleukin‐12 is greater in tobacco hairy roots grown in a mist reactor than in an airlift reactor

Liu

Towler

Medrano

et al. 2008

Biotech & Bioengineering

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We compared the growth and productivity of a tobacco line of hairy roots that produces murine interleukin 12 (mIL-12) grown in three different culture systems: shake flasks, an airlift reactor, and a scalable mist reactor. Of the total mIL-12 produced by cultures grown in shake flasks ( approximately 434.8 microg L(-1)), almost 21% was recovered from the medium. In contrast to roots harvested from shake flasks and the mist reactor, roots were not uniformly distributed in the airlift reactor. Roots formed a dense ring around the wall of the reactor and surrounding the central rising column of fine aeration bubbles. Root quality was also better in both the shake flasks and mist reactor than in the airlift reactor. There were more pockets of dark roots in the airlift reactor suggesting some of the roots were nutrient starved. Although the best root growth (7 g DW L(-1)) was in the shake flasks, both reactors produced about the same, but less dry mass, nearly 5 g DW L(-1). Total mIL-12 concentration was highest in the mist reactor at 5.3 microg g(-1) FW, but productivity, 31 microg g(-1) FW day(-1) was highest in shake flasks. Roots grown in the mist reactor produced about 49.5% more mIL-12 than roots grown in the airlift reactor. Protease activity in the media increased steadily during culture of the roots in all three systems. The comparisons of protease activity, protein and mIL-12 levels done in the shake flask system suggest that the increase in proteases associated with progression into stationary phase is most detrimental to mIL-12 concentration. This is the first description of the design and operation of a scalable version of a mist bioreactor that uses a plastic bag. This also the first report of reasonable production levels of functional mIL-12, or any protein, produced by hairy roots grown in a mist reactor. Results will prove useful for further optimization and scale-up studies of plant-produced therapeutic proteins.

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“…Furthermore, hairy roots have been found to synthesize secondary metabolites at similar or higher levels to those found in whole plants (Flores 1987;Hu and Du 2006;Oksman-Caldentey and Hiltunen 1996;Rhodes et al 1994;Sevón and Oksman-Caldentey 2002) and to have light-guiding properties (Towler et al 2006). In addition to their recombinant protein expression ability, transgenic hairy root cultures exhibit a significantly greater long-time stability than transgenic suspension cultures (Sharp and Doran 2001).…”

Section: Hairy Root Culturesmentioning

confidence: 96%

“…As readily identifiable from the bioreactor name, the droplet size of mist bioreactors is smaller (usually 0.01-10 µm) than the potentially much greater droplet size of spray bioreactors (10-10 3 µm). Finally, the hydrodynamic stress is low in gas-phase bioreactors (Towler et al 2006;Weathers et al 1999;Whitney 1992;Wilson 1997).…”

Section: General Considerationsmentioning

confidence: 97%

Plant Cell-Based Bioprocessing

Eibl

2009

Cell and Tissue Reaction Engineering

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Plant cell-based bioprocessing is the use of plant cell and tissue cultures for the production of biologically active substances (low molecular secondary metabolites and recombinant proteins). The most significant advantage of plant cell culture over the traditionally grown whole wild plant or engineered transgenic plant is the sterile production of metabolites under defined controlled conditions independent of climatic changes and soil conditions, which means that variations in product yield and quality can be better avoided. Furthermore, regulatory requirements such as the cGMP standards, which have to be adhered to in the early stages of pharmaceutical production, are more easily met.Moreover, plant cells are capable of performing complex posttranslational processing, which is a precondition for heterologous protein expression. When compared with mammalian cells, which currently dominate in the commercial protein manufacture, plant cell cultures as alternative expression systems guarantee safer processes because there is a lower risk of contamination by mammalian viruses, pathogens, and toxins. In addition to this considerable advantage, the process costs can also be substantially reduced. This is due to the fact that plant cell culture medium is very simple in composition and therefore relatively inexpensive.This chapter provides an overview of culture types, techniques, and suitable bioreactors used to produce secondary metabolites and recombinant proteins in plant cells. We describe plant cell culture basics, discuss key topics relevant to plant cell bioreactor engineering with application examples, and give an overview of approaches to improving productivity of plant cell-based processes.

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Design, Development, And Applications Of Mist Bioreactors For Micropropagation And Hairy Root Culture

Cited by 8 publications

References 66 publications

Antioxidant Potential and Enhancement of Bioactive Metabolite Production in In Vitro Cultures of Scutellaria lateriflora L. by Biotechnological Methods

Antioxidant Potential and Enhancement of Bioactive Metabolite Production in In Vitro Cultures of Scutellaria lateriflora L. by Biotechnological Methods

Production of mouse interleukin‐12 is greater in tobacco hairy roots grown in a mist reactor than in an airlift reactor

Plant Cell-Based Bioprocessing

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