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Revision as of 21:42, 31 October 2017
DESIGN
The bacteria-robot interface is realized as a highly modular feedback system.
A mobile robot’s sensor values alter the environment of E. coli strains that were designed to respond to these changes with increased expression of fluorescent proteins. In turn, this fluorescence is measured and its quantities are translated into robot behaviour.
For how it all played out, see Results, but let’s first have a glimpse at the modules comprising the feedback loop (for a full description see the different sections in "Parts").
The Bacteria
"ibpA" - Heat Shock Promoter
The heat shock promoter ibpA is controlled by the transcription factor σ 32. In principle, the exposure to high temperatures leads to an increase of σ 32, which subsequently enables heat shock promoters to be recognized by the RNA polymerase. The promoter exhibits a high induction rate and high levels of expression. In our experiment, the ibpA promoter controls the expression of GFP.
"asr" - Acid Inducible Promoter
Promoter activity is controlled by the RstAB System detecting the pH and the PhoRB System activated when inorganic phosphate is rare. Thus, expression only works in low phosphate media (LPM). When grown in LPM and activated by a switch of pH to 5,5 the promoter becomes active and mCardinal is expressed. To enhance expression an extra ribosome binding site (RBS) was inserted between the promoter and mCardinal.
"alx" - Alkaline-induced Roboswitch
Regulation of translation is managed by a pH sensitive riboswitch. The riboswitch itself, a mRNA part 5’ of the RNA coding for our green fluorescence mNeonGreen is regulated by a constitutive promoter.Hence regulation of mNeonGreen translation is managed by the riboswitch in subjected to pH. When pH is neutral the structure of the riboswitch prevents the ribosome from binding to the RBS. When pH rises, structure of the mRNA changes and allows the ribosome to bind the RBS and therefore translation can start.
The Bioreactor
Our system consist of several modules, we differentiate them in 3 layers seen in figure 3. The layer on top is the INPUT layer, it is a steady source of medium or variable liquid, which can be changed and is a key component for its variability. The ANALYSE and MAINTAIN layer consists of two elements and the reactor important to maintain our culture and two separate measuring units, one of which is the D 600 measure unit and the other one can be changed is a variable component (see Interaction Modules (IMs) and Fluorescence Measurement Chamber (FMC)). Both the variable liquid and the variable measure unit gives us the freedom to change our system for different projects. In the OUTPUT layer we collect the waste from our measure units. Further to obtain a steady D 600 value the D 600 measure unit also transports medium containing cell mass to the waste to regulate the D 600 value if it is too high.
Interaction Modules (IMs)
Temperature IM
pH IM
Fluorescence Measurement Chamber (FMC)
Server
Robot
Arena
OVERVIEW fehlt!!!!!