Part:BBa_K2301003

This part contains a temperature dependent promoter designated TlpA36, followed by a hindIII cut site, a sigma 70 constitutive promoter, a ribosome binding site, and, finally, the TlpA36 protein. The hindIII cut site can be used to regulate any gene of interest put into the sequence. TlpA36 is a mutant temperature-dependent promoter/repressor system first outlined in Tunable thermal bioswitches for in vivo control of microbial therapeutics (Piraner et. al. 2016). It is a mutant version of a promoter found naturally in Salmonella typh- Imurium. It strongly represses whatever is downstream of it, except when it undergoes a drastic deformation in a narrow range of temperatures, from 37C to 45C. Inside this range, and only inside this range, will a gene under the promoter’s control be expressed. The repressor protein itself is under the control of a sigma 70 constitutive promoter so that it is always expressed regardless of temperature. It is possible to edit our part to remove the sigma 70 promoter to place the TlpA36 repressor protein under the control of the TlpA36 promoter to create a feedback loop. We incorporated this part along with BBa_K2301001, BBa_K2301002 and BBa_K2301000 into a temperature controlled kill switch for bacteria.

Characterization: For characterization of the TlpA36 repressor system alone, please see Tunable thermal bioswitches for in vivo control of microbial therapeutics (Piraner et. al. 2016). We attempted to characterize the TlpA36 repressor system in conjunction with T4 holin, T4 antiholin, and T4 endolysin as outlined on our project page http://2017.igem.org/Team:Michigan.