Team:Austin UTexas/Improve
Improve
BBa_K2657005: sYFP2 with Mutational Hotspot Removed
BBa_K2657005 is an improved version of the Biobrick BBa_K864100, which is the coding sequence for super yellow fluorescent protein (sYFP2). We increased the evolutionary stability of the sequence by removing the mutational hotspot. Originally, BBa_K864100, contained a palindrommic sequence. Palindromes in the DNA cause potential hairpins, which prevents RNA polymerase from translating the DNA correctly. We then designed primers that added BsmBI and BsaI restriction sites to the sequence and inserted it into PhytoBrick universal acceptor, BBa_P10500, via BsmBI assembly. This created a Phytobrick that functions in Golden Gate Assembly reactions which will be useful due to it's strong fluorescent character.
BBa_K2657003: RCP Phytobrick
In order to improve a part already in the Biobrick Regisry, we chose the Red Chromoprotein-expressing, BBa_E1010. We then designed primers that added BsmBI and BsaI restriction sites to the sequence and inserted it into PhytoBrick universal acceptor, BBa_P10500, via BsmBI assembly. This created a Phytobrick that functions in Golden Gate Assembly reactions.
BBa_K2657004: RCP Phytobrick with a Strong Promoter
For this improvement, we took BBa_E1010 (RCP) and added the synthetic, strong constitutive promoter, CP25. This will increase the expression of the RCP. We also made the sequence golden gate compatible by inserting the sequence into the PhytoBrick Universal Acceptor, BBa_P10500. By linking a Promoter/RBS with a coding sequence, the transformation efficiency of Golden Gate Assembly reactions will increase since the number of parts in the assembly will be reduced. This will also allow BBa_E1010 to be a more useful selective marker as it usually requires one or two days to express and expresses weakly in its current form.
