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Studies on Optimization of the Process of Gellen Producing

Author: ChenYaMin
Tutor: WuXueChang
School: Zhejiang University
Course: Microbiology
Keywords: Sphingomonas elodea gellan fermentation process dissolved oxygen Vitreoscilla hemoglobin vgb
CLC: TQ920.1
Type: Master's thesis
Year: 2011
Downloads: 150
Quote: 0
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Abstract


This study was carried out in an attempt to address the technical problems associated with the process of gellan fermentation production. The bacterium used for the production of gellan is Sphingomonas elodea ATCC 31461 which is Gram-negative, rod-shaped and strictly aerobic. The fermentation medium used to produce gellan is characterized by a high C:N ratio and provides a complex ingredient supplying vitamins (eg., yeast extract, Promsoy) to enhance cell growth. Most reported medium and cultivation condition are not too different, and here our results suggested that the best carbon source was sucrose and the optimum nitrogen source was soy protein flour. Also a 10% inoculum was used for subsequent cultivation in a bioreactor and the cultivation was carried out at 30℃.Then VHb gene was expressed in Sphingomonas elodea ATCC 31461 under the control of constitutive bla promoter to improve oxygen uptake and utilization. Biochemical activity of expressed VHb was confirmed by CO-difference spectra analysis which exhibited a characteristic absorption maximum at 419 nm. During cultivation, not only enhanced cell growth was detected, but also 20% improvement of gellan production was observed after 48 h of incubation in a 6.7L stirred bioreactor, with a maximum yield of 16.82g/L. Moreover, maximum sucrose conversion efficiency (g gellan/g sucrose) was 57.8%,20% higher than that of the parental strain. We further examined the polysaccharide production of VHb-expressing strain at different aeration levels in Erlenmeyer flasks. Again, in all cases a significant enhancement of gellan production was observed, and the enhancement was more significant under oxygen-limiting conditions (up to 26.8%). Thus VHb exhibited positive effect on cell growth and gellan yield of Sphingomonas elodea, especially under hypoxic conditions.This is the first application of VHb as an effective metabolic engineering strategy in Sphingomonas elodea to regulate cell growth and optimize gellan yield. And this can facilitates the commercial developments and applications of gellan gum. Also, our results provided potential applications of VHb in other industrially important strains of the bacterial genus Sphingomonas to solve the increasing demand of aerobic processes in polysaccharides production.

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CLC: > Industrial Technology > Chemical Industry > Other chemical industries > Fermentation industry > General issues > Basic theory
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