Davidson Missouri W/Gene splitting

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=What is Gene Splitting?=

"Gene splitting" refers to the insertion of a hixC site within the coding region of a gene. Although this allows us to create edges for the simulation of a graph, it will change the protein sequence, potentially interfering with proper functionality. We successfully inserted hixC in two different reporter genes, GFP and RFP. Cells transformed with plasmids containing these "split" genes still fluoresce the appropriate color.





To facilitate the splitting process we developed software to help us. Our online gene splitting web tool (click here for a tutorial) helps us choose PCR primers that will amplify the appropriate segments of a gene of interest.

=The Genes=

=A Closer Look=

Here is a more detailed view of splitting GFP. The image on the left, with the protein structure highlighted in green, shows the normal, wild-type structure of the protein. The image on the right highlights the two parts of the protein on either side of the hixC insertion, shown in pink. The additional 13 amino acids are not shown, but they would extend from one pink amino acid into the next.

From these 3-dimensional PDB images it becomes apparent that the insertion likely sticks into open space away from the rest of the protein. This increases the likelihood that the addition does not interfere with the beta barrel structure or with the chromophore.

As a consequence of RFP's similar 3-dimensional structure it is assumed the hixC insertion acts in a similar fashion.



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