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The overall mission of our team is to attempt to overcome some practical barriers to entry of groups and laboratories that may not be well funded or may not have the capital requirements to realize their synthetic biology dreams. Our goal this year is to provide a way for anyone to be able to make their own Taq polymerase instead of having to purchase it. However, the gene for Taq polymerase has a PstI site located within the coding sequence which makes it incompatible with the BioBrick assembly standard. In order to make the Taq polymerase compatible with the BioBrick assembly standard, we need to change the sequence through site directed mutagenesis in a way that would maintain the amino acid sequence of the protein but change the DNA sequence of the gene. In addition, we will be adding the Bio-brick prefix and suffix as well as promoter, terminator and ribosome binding site. Finally, we will be able to generate our own Taq polymerase. We will test the protein to check if our Taq polymerase is active, specific and robust in comparison to the commercial Taq polymerase. If we are successful then this polymerase will reduce the cost of the one of the most expensive reagents in synthetic biology.

Project Details

While we designed the point-mutagenesis primers, we took the opportunity to design and order the primers for the BioBrick Suffix and Prefix. We followed the examples laid out in the Registry of Standard Parts for designing the oligos needed to make a part. Important considerations are melting point and CG concentration, as well as self-dimerizations and hairpins. We analyzed these primers using the OligoAnalyzer from IDT. When analyzing PolI, only the coding seuence itself was used for sequence inquiry, not the BioBrick Suffix/Prefixes.