0102 Synthetic Biology Notes
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Transcript of 0102 Synthetic Biology Notes
Synthetic Biology Notes
General Terms
• Ex vitro
In vivo (“within the living)– experimentation using a whole, living organism (e.g. animal testing, clinical trials) ex vivo (“out of the living”) –
experimentation/measurements done in or on tissue in an artificial environment outside the organism with the minimum alteration of natural conditions
In vitro (“within the glass”) – experimentation using a controlled environment (ex. Test tube/petri dish)
in silico– performed on computer or via computer simulation
A SYNTHETIC GENETIC EDGE DETECTION PROGRAM
Tabor 2009
System• Mask filters light detected by bacteria• bacteria/ lawn detects edges + gives visual output
Concept of Detection System• Bacteria in dark produce signal (green circle)• Signal diffuses• Only bacteria in light respond to signal;
• Response = expression of visible reporter gene
NOT light box
cell-cell communication
inverter
X AND (NOT Y)
• light, no signal, no pigment• light, signal, pigment• No light, no signal, no pigment• no light, signal, no pigment
IF NOT lightproduce signal;
IF signal AND NOT (NOT light)produce pigment;
Y = 0 when there’s light.
light sensitive protein
cph8 = [Cph1]+[EnvZ]• Cph1 = photoreceptor domain of Synechocystis• EnvZ = kinase domain of E.coli • Inhibited with red light• Requires phycocyanobilin (PCB)
• Covalently associated chromophore• Produced from heme by genes ho1 and
pcyA
PCB
resp. reg.OmpRphorphoryl
group
Sensor• Phytochrome– Membrane bound extracellular sensor/photoreceptor– Intracellular (response) regulator• Usu do not have DNA-binding domains• Do not directly regulate gene expression
• Fusion protein: phytochrome and EnvZ histidine kinase domain
Extracellular sensor
Intracellular regulator
Photoreceptor Expression
• PCB not produced in E.Coli• Introduction of PCB synthesis genes• Converts haem into PCB
Transduction Mechanism
• Red light represses EnvZ auto-phosphorylation
Output
• OmpR-P activates promoter linked to reporter gene
• lacZ reporter produce black compound
OmpR-P dependent ompC promoter
System Revisited
Light Canon
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