Endy:Chassis engineering/Orthogonal protein synthesis
Dedicated systems
- Dedicated systems are a method to decouple the function of an engineered biological system from the function of its chassis.
- By separating the resources and machinery used to supply and power an engineered system from those of the chassis, then perturbations in the operation of one should have less effect on the other.
- Currently, we are working to implement these systems at a transcription and translation level.
- These dedicated synthesis systems can then be used as the basis of Biological virtual machines
- The different methods we are using to implement dedicated systems are described in more detail below. We have built reporter devices to examine the performance of the four different combinations of transcription systems currently available to us. They are shown in matrix form below.
Reporter devices for the transcription/translation systems
| General Translation | VM Translation | |
|---|---|---|
| General Transcription | I7101 |
I7102
|
| VM Transcription | E7104 |
E7103
|
VM Transcription
Introduction
The T7 expression system developed by Studier and coworkers is essentially orthogonal from the E. coli transcription system. T7 RNAP does not recognize E. coli promoters and E. coli RNAP does not recognize T7 promoters.
Effects of producing dedicated transcription machinery in a cellular chassis
Three colonies of BL21(DE3) were grown in the presence or absence of 0.4mM IPTG. The IPTG induces the production of T7 RNAP. Results suggest that the presence of dedicated transcription machinery in BL21 does not noticeably affect the growth rate of the cellular chassis.
While this implies that the presence of a dedicated transcription system has negligible effect on a cells ability to supply a demand, it is possible that the added demand of these systems pushs the cellular chassis much closer to its maximum demand level before physiological changes become evident.
VM Translation
