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Coming up with a project
Teach the Teachers WorkshopMay 16, 2009
Talk to people
Previous iGEM projectsigem.org
New organisms
New parts and tools for future teams
Most commonly used parts: B0015 - a terminator
F2620 - an inducible promoterB0034 - a RBS
R0011 - lac promoterPlasmid backbones
Reuse and existing parts
Mechanism & Function
A transcription factor (LuxR) that is active in the presence
of a cell-cell signaling molecule (3OC6HSL) is controlled by
a regulated operator (PLtetO-1). Device input is 3OC6HSL.
Device output is PoPS from a LuxR-regulated operator. If
used in a cell containing TetR then a second input such as
aTc can be used to produce a Boolean AND function.
BBa_F26203OC6HSL PoPS Receiver
Registry of Standard Biological Parts making life better, one part at a time
License: Public
Conditions (abridged)
Output: PoPS measured via BBa_E0240
Culture: Supplemented M9, 37ºC
Plasmid: pSB3K3
Chassis: MG1655
*Equipment: PE Victor3 multi-well fluorimeter
**Equipment: BD FACScan cytometer
1E01E1
1E21E3
1E4
GFP (arbitrary units)
Doublin
gs
20
38
56
74
92
High Input (1E -7 M 3OC6HSL)
92
1E01E1
1E21E3
1E4
Doublin
gs
20
38
56
74
Low Input(0 M 3OC6HSL)
GFP (arbitrary units)
Reliability**
Genetic: >92/>56 culture doublingsPerformance: >92/>56 culture doublings
(low/high input during propagation)
Transcriptional Output Demand (low/high input)
Nucleotides: 0 / 6xNt nucleotides cell-1 s-1
Polymerases: 0 / 1.5E-1xNt RNAP cell-1
(Nt = downstream transcript length)
Chassis: MC4100, MG1655, and DH5
Plasmids: pSB3K3 and pSB1A2
Devices: E0240, E0430 and E0434
BBa_F2620 Response Time: <1 minBBa_T9002 Response Time: 6±1 minInputs: 0 M (Low), 1E-07 M (High) 3OC6HSL
Pmax: 6.6 PoPS cell-1
K: 1.5E-09 M 3OC6HSLn: 1.6
Static Performance*
Pout =Pmax [3OC6HSL]n
Kn+ [3OC6HSL]n
0E+00 1E!10 1E!09 1E!08 1E!07 1E!06 1E!05 1E!04
0
100
200
300
400
500
600
[3OC6HSL] (M)
GF
P s
yn
the
sis
ra
te (
mo
lec
ule
s c
ell!1 s!1)
Population Mean
Colony Range
Hill Equation
0
1
2
3
4
5
6
7
8
Po
PS
ce
ll!1
Part Compatibility (qualitative)
Authors: Barry Canton Ania Labno
Updated: March 2008
Sig
nalin
g D
evic
es
http://p
art
s.m
it.e
du/r
egis
try/index.p
hp/P
art
:BB
a_F
2620
BBa_F2620
R0040 B0034 C0062 B0015 R0062 PLtetO-1 RBS luxR Term. Plux,R
Component Parts
!10 0 10 20 30 40 50
0
100
200
300
400
500
600
+ 3OC6HSL
Time (min)
GF
P s
yn
thesis
rate
(m
ole
cu
les c
ell!1 s!1)
GFP synthesis rate (Low Input)
GFP synthesis rate (High Input)
Polynomial Fit (High Input)
PoPS (High Input)
0
1
2
3
4
5
6
7
8
Po
PS
cell!1
0E+00 1E!10 1E!09 1E!08 1E!07 1E!06 1E!05 1E!04
0
100
200
300
400
500
600
[AHL] (M)
GF
P s
yn
the
sis
ra
te (
mo
lec
ule
s c
ell!1 s!1)
C4HSL
C6HSL
3OC6HSL
C7HSL
C8HSL
3OC8HSL
C10
HSL
C12
HSL
0
1
2
3
4
5
6
7
8
Po
PS
ce
ll!1
Input Compatibility*
Dynamic Performance*
Let the students choose
Help them make smart choices
• Figure out what’s practical: How many assembly stages could the team possibly do over the course of the summer? That sets an upper limit to the size of the system.
• Design the project so that different modules can be done in parallel.
• It doesn’t have to be a brand new idea.
Describe your project on your team wiki
Teach the Teachers WorkshopMay 16, 2009
Standard assembly
Teach the Teachers WorkshopMay 16, 2009
BioBrick standard parts
EcoRI XbaI SpeI PstIBioBrick part
Knight, 2003
BioBrick standard assembly
E X S PBioBrick part 1 E X S PBioBrick part 2
Digest with EcoRI and SpeI
Digest with XbaI and PstI
E X SBioBrick part 1 E X S PBioBrick part 2
E X S PBioBrick part 1 BioBrick part 2Mixed
Ligate
A
B
C
D
E
F
G
H
AB
CD
EF
GH
ABCD
EFGH
Why use the BioBrick standard?
• It is faster to build multi-part systems
• Assembling every two parts is the same
• You can reuse parts from the Registry
• Other people can reuse your parts
• It is required to win a prize at iGEM!
Get the enzymes cheaper
Plasmid backbones
Teach the Teachers WorkshopMay 16, 2009
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