Team:Purdue/Genetic Pathway
Background Information
There are two major techniques we apply in our experiment design. One of them is the usage of split proteins. These are proteins whose coding regions have been split in half in thier DNA in order to create a binary effect by producing 2 different protein halves. These are used to allow protein-protein interactions. In our case, we have our sensing protein being split that is also bound to a split reporter molecule, in our case it will be HRP. According to ...'s research, HRP was split in various locations to serve as a lock mechanism. However, the ideal spot to split the protein wasn't known at the time. Instead, they did a process called rational design where they analyzed the protein to find sites where this process would most likely succeed. They then created different variations where they tested their mechanism at these various sites. Using their findings, we are using the 58 site for the HRP split.
However, unlike what the paper did, we didn't/don't know if our locking mechanism will work. So to do this, we needed a way to find how to split our sensing protein at sites that worked for our purposes. Since, most of our team lacks the training and the background involving protein structure and mechanics. Therefore, we are doing irrational design. This is a process where we generate various different split sequences. The procedure we used is similar to what is found in this paper here
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Experiments
Describe the research, experiments, and protocols you used in your iGEM project. These should be detailed enough for another team to repeat your experiments.
Please remember to put all characterization and measurement data for your parts on the corresponding Registry part pages.