November 10, 2015 at 5:59 am #18181chanceParticipant
The E. coli expression system is extensively used as the first choice in genetic engineering due to its high productivity, high growth and production rate, ease of use, and economy, etc. However when expressed in e.coli, most recombinant proteins deposit in the form of insoluble and inactive inclusion bodies.In vitro renaturation is a complicated and low-efficient process.
The purpose of the invention can be achieved by the following method, which comprises introducing the leader sequence at 5′-UTR (untranslatable region) of tobacco mosaic virus (TMV) genome into an E. coli expression vector, between the promoter region and the ribosome binding site (rbs), thereby forming a novel expression vector which increases the yield of soluble, biologically active recombinant products and effectively avoids or improves the formation of inclusion bodies of recombinant proteins.
In the first aspect, the invention provides an E. coli expression vector comprising an omega leader sequence between the promoter region and ribosome binding site.
In a preferred embodiment, the omega leader sequence comprises the sequence of SEQ ID NO: 2.
In a preferred embodiment, said expression vector comprises a core regulatory region having the sequence of SEQ ID NO: 1.
In a preferred embodiment, said expression vector is a GST fusion expression vector.
In the second aspect, the invention provides an E. coli comprising the E. coli expression vector of the invention.
In the second aspect, the invention provides a method for improving an E. coli expression vector comprising the steps of:
1. providing an E. coli expression vector which comprises an promoter region and ribosome binding site;
2. inserting an omega leader sequence between said promoter region and ribosome binding site, thereby forming an expression vector which comprises said omega leader sequence.
In a preferred embodiment, said method further comprises the step of:
3. inserting the GST coding sequence, the multiple cloning sites, and the histidine tag coding region orderly into said vector, downstream of the ribosome binding site, thereby forming a GST fusion expression vector.
For more detailed information about enhancing solubility of protein expression in E. coli, you can refer to this PDF file：http://www.biotechniques.com/multimedia/archive/00036/BTN_A_04373ST07_O_36989a.pdf
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