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Library of Plasmids to Control Gene Expression Levels
EIB-2792

APPLICATIONS OF TECHNOLOGY:

ADVANTAGES:

ABSTRACT:

Taek Soon Lee and colleagues at the Joint BioEnergy Institute (JBEI) have developed a technology that can be used to control the expression of different genes transformed into host cells. The scientists constructed a library of 96 expression vectors with various combinations of replication origins, transcriptional promoters, and antibiotic resistance markers, the first two of which control gene expression level. These vectors are also listed in a database. A scientist wishing to differentially express a set of genes can query the database, choose the most appropriate vectors for the desired expression levels, and then splice each gene of interest into the appropriate vector using BglBricks, a robust splicing technology. When the plasmids are transformed into host cells, the gene expression levels may be fine-tuned by adjusting the level of appropriate inducers.

After initially creating the expression vectors, the JBEI scientists spliced into them red and green fluorescent proteins whose expression levels could be quantified for creation of the database. They then used appropriate vectors to increase the expression levels of two enzymes that were bottlenecks in a pathway involved in biofuel production. This successfully led to not only elevated expression of the enzymes but increased productivity of the pathway—an important step in improving biofuel technology.

In metabolic engineering and synthetic biology, researchers identify valuable enzymatic pathways, such as those that produce medications, break down toxic wastes, or convert inexpensive plant material into biofuel. To engineer these pathways (place them in host cells and control the composite reactions), one must be able to influence the expression of the genes that code for the constitutive enzymes. This has been a significant challenge in genetic engineering. The JBEI library of expression vectors overcomes this challenge by allowing scientists to efficiently place target genes in commonly used host cells and control their expression.

DEVELOPMENT STAGE: Industrial prototype.

STATUS: Available for licensing or collaborative research.

FOR MORE INFORMATION:

Lee, T.S., Krupa, R.A., Zhang, F., Hajimorad, M., Holtz, W.J., Prasad, N., Lee, S.K., Keasling, J.D. “BglBrick Vectors and Datasheets; a synthetic biology platform for gene expression,” Journal of Biological Engineering, Sept. 20, 2011.

A.M. Redding-Johanson, Batth T.S., Chan R., Krupa R., Szmidt H.L., Adams P.D., Keasling J.D., Lee T.S., Mukhopadhyay A., Petzold C.J. “Targeted proteomics for metabolic pathway optimization: application to terpene production,” Metab Eng. 13 (2011) 194-203.

SEE THESE OTHER BERKELEY LAB TECHNOLOGIES IN THIS FIELD:

j5: Automated DNA Assembly Design Software, ECRB-2836

Directed Evolution of Microbe Producing Biofuels Using In Vivo Transcription Factor Based Biosensors, EJIB-2710

Irreversible Low Load Genetic Switches, EJIB-2593

REFERENCE NUMBER: EIB-2792

 

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