Part:BBa_K5101005
pET29(a)-pJ23119-TIpA39-T-pTIpA-RBS-PhiX174E-T7
temperature-inducible suicide module
Usage and Biology
To enable the engineered bacteria to suicide when released into the environment (generally above 39 degrees Celsius) during fecal disposal, we chose a temperature-responsive protein TlpA39(BBa_K2572000) and its corresponding promoter pTlpA(BBa_K2500003) as regulatory elements to induce the expression of the lysis protein PhiX174E. TlpA39 is derived from Salmonella enterica. It is a temperature-sensitive protein that can bind to pTlpA. At temperatures above 39 degrees Celsius, the TlpA39 protein will remain in a monomeric form, thereby losing the ability to bind to the promoter pTlpA. If the temperature drops below 39 degrees Celsius, the TlpA39 protein will form a dimer; the dimer can inhibit the pTlpA and prevent the recruitment of RNA polymerase, thereby suppressing downstream gene expression.
Design
As shown in Figure 1, we designed the plasmid pET29(a)-pJ23119-TIpA39-T-pTIpA-RBS-PhiX174E-T7 using SnapGene software. Through the continuous expression of the TIpA39 protein under the strong constitutive promoter J23119, the pTIpA promoter is inhibited at temperatures below 39 degrees Celsius. When animal manure is being processed and the temperature reaches above 39 degrees Celsius, it will initiate the expression of the lysis gene, achieving the self-destructive effect. In the future, we will complete wet experimental validation of this suicide module.
Figure 1 Plasmid pET29(a)-pJ23119-TIpA39-T-pTIpA-RBS-PhiX174E-T7
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30
Illegal NheI site found at 1205
Illegal NotI site found at 572 - 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 857
Illegal XhoI site found at 1632 - 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1160
- 1000COMPATIBLE WITH RFC[1000]
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