Construction of Gene Expression Cassette
Introduction
Gene expression cassette in viral vectors can be complex. Basic expression cassette has a eukaryotic promoter, open reading frame of the gene-of-interest, and a poly-A signal sequence. However, one can add elements such as intron, 3'-UTR, regulatory elements to enhance gene expression or confer specific gene expression profile in target cells. 

Another level of complexity can be added when the researcher wants to express more than one gene from the same viral vector. Frequently a marker gene such as GFP is co-expressed together with the gene-of-interest to serve as indicator of successful transduction of target cells. 

Further more, siRNA expression, regulated gene expression (on/off control), Cre-loxP system all can be integrated into viral vector. These advanced features may need more than one viral vector to accomplish. Developing such a viral vector is a big challenge. 

 
What we can do
Applied Viromics offers shuttle plasmids that contain basic gene expression cassette, such as a strong promoter (such as CMV, CAG), a multi-cloning site for subcloning of trans-gene, and a poly-A. The construct is extensively tested and proven to work well.

Options are available to express two genes from the same vector. An ires element can be used to translate two proteins from one mRNA, or two genes can be transcribed from two promoters.

We also have special designed plasmids to express siRNA from mU6 promoter. siRNA can be expressed alone, or with a marker gene.

Some expression-enhancing elements are available, such as introns, 3'UTRs, for consideration when designing an expression cassette. What we will do at Applied Viromics is to help you come up with a design that could deliver the result you want, and offer the service to prepare the plasmid for you to use in viral vector production.

 
Service detail
We will work together with investigators aiming to integrate our in-house viral plasmid with whatever available from investigator. 

First, we offer consulting service to help our customer to finalize a design of the viral vector expression cassette, and access the availability of all elements needed to construct the viral plasmid.

The shuttle plasmids we have at Applied Viromics are available for investigators to subclone their expression cassette without charge, given that we perform the viral production service using those plasmids.

When new plasmids need to be constructed to accommodate the needs of the investigator, it is the choice of the investigator to do the job themselves, or use our service. Our service charge is 500$ for each step of subcloning. Depending on the complexity of the project, number of subcloning steps necessary to make the new plasmid is calculated and a quotation will be submitted to customer.

Applied Viromcis will be responsible to the integrity of the plasmid we constructed, by means of restriction enzyme digestion or/and sequencing at a reasonable level. It is the customer's responsibility to test the functionality of the construct.

 
Terms and conditions
Customer owns the plasmids prepared for their project. Applied Viromics will release all material obtained from the customer for making the new plasmid together with the final viral plasmid.

Customer shall make available of the "building blocks" needed to construct the viral plasmid they request when needed.

Customer shall obtain user's right for all patent-protected material used in their project. Applied Viromics does not provide such material nor offer any license to our customer. Applied Viromics will not be hold responsible for any misuse of patented material in customers viral plasmids. It is the customers responsibility to make sure they have all the right to use any material in their project.