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Gene Design 2 - Gene Constructs

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Roundup Ready

EPSPS or 5-enolpyruvyl-shikimate synthase is an enzyme that catalyzes a reaction in plant and bacterial cells that is necessary for the synthesis of some amino acids. The herbicide, Roundup, can bind to the EPSPS enzyme made in plants and block its ability to work. This causes the plant to run out of amino acids which halts growth and development. The plant eventually dies from starvation. Roundup is a non-selective systemic herbicide which means it injures and often kills most plants that come into contact with it.

 The version of EPSPS made in a special strain of Agrobacteria has a slightly altered shape. This alteration prevents Roundup from binding, thus allowing the resistant EPSPS to catalyze the amino acid synthesis reaction. The coding region from the bacterial EPSPS gene was cloned and transferred to plants. Plants with the bacterial EPSPS can thus be sprayed with Roundup and take it up into their cells. Now, they have a backup enzyme that gives them the ability to keep making amino acids and thus resistance to Roundup.

The backup enzyme can be compared to a detour route around road construction. Cars are still able to get to their destination, but the traffic moves slower. In the same way, the backup enzyme provides an alternate pathway for amino acid synthesis to continue. However, the pathway may not be as efficient and thus unable to produce as many amino acids. This can result in a slight decrease in yield especially if Roundup is applied at high rates.

The promoter used in the Roundup Ready gene is the 35S promoter. Thus the gene is expressed in all plant cells. A further modification in the transgene is the addition of sequences to the coding region which places additional amino acids on the end of the EPSPS enzyme. These additional sequences are called the chloroplast transit peptide (CTP). They direct the EPSPS protein to go to the chloroplasts in the plant cell.
The Roundup Ready gene uses the 35S promoter to give expression in all cells. It also has the EPSPS (Roundup resistance) coding region with the addition of the CTP coding segment to direct the EPSPS protein to the chloroplasts.

EPSPS is one of the many proteins made in plants that need to go to the chloroplast to get their work done. Agrobacterium doesn’t have chloroplasts and consequently does not have this sequence on its gene for EPSPS. If the CTP region is not added to the transgene before transformation into a plant, the Roundup resistant EPSPS protein will not get to the parts of the plant that need it most which are the chloroplasts found in stems and leaves.


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