Description

The Tse2 protein is a toxin component of a toxin-immunity system and arrests the growth of prokaryotic and eukaryotic cells when expressed intracellularly. In contrast, secreted Tse2 has no effect on eukaryotic cell.

Assembly

The BioBrick BBa_K314200 coding for the Tse2 toxin was assembled with the weak BBa_B0031 RBS and with the BBa_B0015 terminator. This composite was then cloned under the inducible promoter T5 LacO that we submitted as BioBrick BBa_K875002 in order to be able to follow the toxin activity when expressed in E.coli

Results

M15 bacteria cells (containing the pREP4 plasmid coding for the Lac repressor) are transformed with this Biobrick that resides on pSB1C3 plasmid.

After an over night liquid culture (LB broth) the OD600 absorbance is recorded and the culture is diluted to 0,1 and the batch is splitted into 6 flask – 20ml each. Three of these are induced with IPTG 1mM and the others are not induced.

The OD 600 is then recorded each hour.

It is possible to observe that not induced cells grow up to OD 1, instead the induced ones do not growth during the time, demonstrating the inducible promoter and toxin efficiency.

Looking forward

In the final system, the toxin should be under the control of the T5 Cumate Operator (BBa_K875001).

Link to the Registry