Ghent University Academic Bibliography

Advanced

A constitutive expression system for high-throughput screening

Dirk Aerts UGent, Tom Verhaeghe UGent, Marjan De Mey UGent, Tom Desmet UGent and Wim Soetaert UGent (2011) ENGINEERING IN LIFE SCIENCES. 11(1). p.10-19
abstract
To reduce the amount of consumables and number of pipetting steps in high-throughput screening, a constitutive expression system was developed that comprises four different promoters of varying strength. The system was validated by the expression of different sucrose phosphorylase enzymes from Leuconostoc mesenteroides, Lactobacillus acidophilus and Bifidobacterium adolescentis in 96-deep-and low-well plates at three temperatures. Drastically improved soluble expression in mini-cultures was observed for the enzymes from L. mesenteroides strains by reducing the promoter strength from strong to intermediate and by expressing the proteins at lower temperatures. In contrast, the enzymes from B. adolescentis and L. acidophilus were expressed most efficiently with a strong promoter. The constitutive expression of sucrose phosphorylases in low-well plates resulted in a level of activity that is equal or even better than what was achieved by inducible expression. Therefore, our plasmid set with varying constitutive promoters will be an indispensable tool to optimize enzyme expression for high-throughput screening.
Please use this url to cite or link to this publication:
author
organization
year
type
journalArticle (original)
publication status
published
subject
keyword
MICROTITER PLATES, LEUCONOSTOC-MESENTEROIDES, DIRECTED EVOLUTION, ESCHERICHIA-COLI, RECOMBINANT SUCROSE PHOSPHORYLASE, SOLUBLE EXPRESSION, PROTEIN EXPRESSION, BATCH CULTURE, PROMOTER, GENE, Constitutive expression, Directed evolution, High-throughput screening, Promoter, Sucrose phosphorylase
journal title
ENGINEERING IN LIFE SCIENCES
Eng. Life Sci.
volume
11
issue
1
pages
10 - 19
Web of Science type
Article
Web of Science id
000287671800002
JCR category
BIOTECHNOLOGY & APPLIED MICROBIOLOGY
JCR impact factor
1.925 (2011)
JCR rank
87/157 (2011)
JCR quartile
3 (2011)
ISSN
1618-0240
DOI
10.1002/elsc.201000065
project
Biotechnology for a sustainable economy (Bio-Economy)
language
English
UGent publication?
yes
classification
A1
copyright statement
I have transferred the copyright for this publication to the publisher
id
1209010
handle
http://hdl.handle.net/1854/LU-1209010
date created
2011-04-14 14:37:17
date last changed
2013-02-27 09:42:20
@article{1209010,
  abstract     = {To reduce the amount of consumables and number of pipetting steps in high-throughput screening, a constitutive expression system was developed that comprises four different promoters of varying strength. The system was validated by the expression of different sucrose phosphorylase enzymes from Leuconostoc mesenteroides, Lactobacillus acidophilus and Bifidobacterium adolescentis in 96-deep-and low-well plates at three temperatures. Drastically improved soluble expression in mini-cultures was observed for the enzymes from L. mesenteroides strains by reducing the promoter strength from strong to intermediate and by expressing the proteins at lower temperatures. In contrast, the enzymes from B. adolescentis and L. acidophilus were expressed most efficiently with a strong promoter. The constitutive expression of sucrose phosphorylases in low-well plates resulted in a level of activity that is equal or even better than what was achieved by inducible expression. Therefore, our plasmid set with varying constitutive promoters will be an indispensable tool to optimize enzyme expression for high-throughput screening.},
  author       = {Aerts, Dirk and Verhaeghe, Tom and De Mey, Marjan and Desmet, Tom and Soetaert, Wim},
  issn         = {1618-0240},
  journal      = {ENGINEERING IN LIFE SCIENCES},
  keyword      = {MICROTITER PLATES,LEUCONOSTOC-MESENTEROIDES,DIRECTED EVOLUTION,ESCHERICHIA-COLI,RECOMBINANT SUCROSE PHOSPHORYLASE,SOLUBLE EXPRESSION,PROTEIN EXPRESSION,BATCH CULTURE,PROMOTER,GENE,Constitutive expression,Directed evolution,High-throughput screening,Promoter,Sucrose phosphorylase},
  language     = {eng},
  number       = {1},
  pages        = {10--19},
  title        = {A constitutive expression system for high-throughput screening},
  url          = {http://dx.doi.org/10.1002/elsc.201000065},
  volume       = {11},
  year         = {2011},
}

Chicago
Aerts, Dirk, Tom Verhaeghe, Marjan De Mey, Tom Desmet, and Wim Soetaert. 2011. “A Constitutive Expression System for High-throughput Screening.” Engineering in Life Sciences 11 (1): 10–19.
APA
Aerts, Dirk, Verhaeghe, T., De Mey, M., Desmet, T., & Soetaert, W. (2011). A constitutive expression system for high-throughput screening. ENGINEERING IN LIFE SCIENCES, 11(1), 10–19.
Vancouver
1.
Aerts D, Verhaeghe T, De Mey M, Desmet T, Soetaert W. A constitutive expression system for high-throughput screening. ENGINEERING IN LIFE SCIENCES. 2011;11(1):10–9.
MLA
Aerts, Dirk, Tom Verhaeghe, Marjan De Mey, et al. “A Constitutive Expression System for High-throughput Screening.” ENGINEERING IN LIFE SCIENCES 11.1 (2011): 10–19. Print.