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Evaluation of the GFP signal and its aptitude for novel on-line monitoring strategies of recombinant fermentation processes.

Abstract：

:A high number of economically important recombinant proteins are produced in Escherichia coli based host/vector systems. The major obstacle for improving current processes is a lack of appropriate on-line in situ methods for the monitoring of metabolic burden and critical state variables. Here, a pre-evaluation of the reporter green fluorescent protein (GFP) was undertaken to assess its use as a reporter of stress associated promoter regulation. The investigation of GFP and its blue fluorescent variant BFP was done in model fermentations using E. coli HMS 174(DE3)/pET11 aGFPmut3.1 and E. coli HMS174(DE3)/pET1aBFP host/vector systems cultured in fed-batch and chemostat regime. Our results prove the suitability of the fluorescent reporter proteins for the design of new strategies of on-line bioprocess monitoring. GFPmut3.1 variant can be detected after a short lag-phase of only 10 min, it shows a high fluorescence yield in relation to the amount of reporter protein, a good signal to noise ratio and a low detection limit. The fluorescence-signal and the amount of fluorescent protein, determined by ELISA, showed a close correlation in all fermentations performed. A combination of reporter technology with state of the art sensors helps to develop new strategies for efficient on-line monitoring needed for industrial process optimisation. The development of efficient monitoring will contribute to advanced control of recombinant protein production and accelerate the development of optimised production processes.

journal_name

J Biotechnol

journal_title

Journal of biotechnology

authors

Reischer H,Schotola I,Striedner G,Pötschacher F,Bayer K

doi

10.1016/j.jbiotec.2003.11.007

subject

Has Abstract

pub_date

2004-03-04 00:00:00

pages

115-25

issue

2

eissn

0168-1656

issn

1873-4863

pii

S0168-1656(03)00318-3

journal_volume

108

pub_type

杂志文章

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