Our aim is to circularize the TAL-effector so the proteins are solid during the purification. To do this we use the vector construct of the iGem team of Heidelberg in 2014 that has an Intein mechanism (more information at http://2014.igem.org/Team:Heidelberg). However, we changed a part of the Intein sequence because we wanted to optimize and stabilize the circulation. Another change was made on the repeat side of the TALE- we did into the vector of the team in Heidelberg. If you want to read more about our project concept click here Project.
We designed four circularized vectors with the help of the Heidelberg team’s mechanism and our own linker. All of them contain the TAL-effector sequence, the Strep Tag, the TEV side and the Intein sequence, a special TAL-effector sequence. This is so that they can bind to another DNA sequence. Additionally, two of them contain a GFP sequence.
|Part Number||Type||Part Name||Short description||Length|
|BBa_K2075000||Tag||Strep XT Tag||Strep Tag for protein purification and detection||90 bp|
|BBa_K2075001||Coding||eGFP||enhanced green fluorescence protein, used as reporter protein||717 bp|
|BBa_K2075003||Protein domain||TEV Cleavage Site||encodes a amino acis sequencen, which is a specific cleavage site for the TEV protease||21 bp|
|BBa_K2075004||Plasmid||Vector iGem_02_Ax7L-DS||Combination of TAL effector Ax7L-DS with TEV Site, Intein and Strep tag||5841 bp|
|BBa_K2075005||Plasmid||Vector iGem_02_Hax3-2xNG||Combination of TAL effector Hax3-2xNG with TEV Site, Intein, eGFP and Strep Tag||5396 bp|
|BBa_K2075007||Plasmid||Vector iGem_02_Ax7R-RR||Combination of TAL effector Ax7R-RR with TEV Site, Intein and Strep tag||5739 bp|
|BBa_K2075009||Coding||Hax3-2xNN||TAL-effector protein, binds the DNA sequence: A C A C C C G/A G/A N C A||1777 bp|
|BBa_K2075010||Coding||Intein and linker (N-Term)||useful by the circularization of a protein by ligating the N- and C-terminal ends||524 bp|
|BBa_K2075020||Coding||Ax7L-DS||TAL-effector protein, binds the DNA sequence: A G/A C A C T A T A T A A A C C C C C||3210 bp|
|BBa_K2075021||Coding||Ax7R-RR||TAL-effector protein, binds the DNA sequence: T G/A A A G/A G/A C T T G/A A T G/A A G/A C T||2946 bp|
|BBa_K2075022||Plasmid||Vector iGem_02_Hax3-2xNN||Combination of TAL effector Hax3-2xNN with TEV Site, Intein, eGFP and Strep Tag||5396 bp|
|BBa_K2075023||Coding||Hax3-2xNG||TAL-effector protein, binds the DNA sequence: A C A C C C T T N C A T||1803 bp|
|BB_aK2075024||Coding||Intein and linker (C-Term)||useful by the circularization of a protein by ligating the N- and C-terminal ends||336 bp|