Rewirable circuit - Main Page - 2014.igem.org

Rewirable circuitAn elegant means of system integration

iGEM HZAU-China!October 31, 2014

Disordered parts

TT

TT T

Duplicated the existing parts

Overlapping module

TT T

TT

Natural biological system

Standard parts

Logic elements

Small modules

Synthetic systems vs Natural systems

Synthetic biological system

Background

BackgroundAdaptability—what synthetic biological systems lack

One module, one function.

How to process different information with one module?

• Host overload

• Crosstalk

As the complexity increases

Intuitive solution: Combining circuitsBackground

Our Solution: The rewirable circuit

What if we can alter regulatory pathways and reuse existing parts?

Rewirable circuit

Our approach

Our approachAdvantages

• Eliminating crosstalk between processing modules.

More adaptive with fewer parts used.!Less is more!

• Choosing specific processing modules when needed

• Reducing workload

Viability of our design

Repressilator

T TT lacI cItetR T TlacI cI

Toggle switchlox66lox71

T TT lacI cItetR

Our rewirable circuit

Information processing

lox66lox71

T TT lacI cItetR

Our rewirable circuit

Our Biobricks:BBa_K1368000! BBa_K1368002

Viability of our designInformation processing

Preliminary works

Parameter Valueβ0 0.03

Kdp 0.0069Kdm 0.347Ktl 6.93··· ···

Data collection Characterization

Information processing

Modeling

d[BmRNA]dt

= β0 +βKn

K n + An− Kdm[BmRNA]

d[B]dt

= Ktl[BmRNA]− Kdp[B]

Parameter scanning and dynamic analysis

Limit cycle attractor

Fixed point attractor

An attracting limit cycle emerges as the promoter strength increases.Strength of promoter AStrength of promoter B

Stre

ngth

of p

rom

oter

C

Information processing

Preconditions to implement both oscillation and bistability

• To implement oscillation

• To implement bistability

The weakest promoter must be strong enough.

Two promoters’ strength must be approximately equal.

Information processing

Simulation resultsInformation processing

Another rewirable circuit design

luxpRluxpL

T TT luxI luxRaiiAluxpLluxpR

Our Biobricks:BBa_K1368004! BBa_K1368005! BBa_K1368006

AHL

Information processing

Results from fluorescence microscope

Positive feedback stage Negative feedback stage

TGFPluxpR

LuxI

LuxR-­‐AHL  complex

AiiA LuxI

LuxR-­‐AHL  complex

AiiA

Information processing

Results:experimental results were consistent with our expectationsResults from multifunctional microplate reader

Information processing

Information processingDesign procedures

• Design fitness functions

• Scan the topological space

A

BC

Topology

0

@0 0 �1�1 0 00 �1 0

1

A

A B CABC

Matrix

FitnessA=0.7891

FitnessB=0.8915…

• Match topological structures

• Sort and select

More details can be found in our modeling part

Potential applications

• Living therapeutics

• Environment improvement

• Circuit design in synthetic biology

Information processing

• …

Plenty of time for comparisonLeave it to nature

Information processingPotential applications

• Living therapeutics

• Environment improvement

• Circuit design in synthetic biology

• …

Maintain homeostasisMake next decision

Information processingPotential applications

• Living therapeutics

• Environment improvement

• Circuit design in synthetic biology

• …

Positive feedback: !increase efficiency

Negative feedback:!maintain the lower steady state

PollutionFunction!proteins

Information processingPotential applications

• Living therapeutics

• Environment improvement

• Circuit design in synthetic biology

• …

Potential applications

• Living therapeutics

• Environment improvement

• Circuit design in synthetic biology

Information processing

• …

Improvements in I/O modules Lower the leakage of recombinase

Our Biobricks:BBa_K1368007! BBa_K1368009! BBa_K1368013!BBa_K1368015

Tcre

Tcre

Problem: leakage

Input

taRNA

cre

Solution: riboregulator

Riboregulator assay

1 !71

1.9 Fold 2.5 Fold 5.5 Fold

Positive control

Negative control

Experimental group

Improvements in I/O modules

Increase response rates in output module

FluorophoreRNA aptamer

+

Fluorescent protein

Improvements in I/O modules

RNA level fluorescence

DMHBI DMHBI + !13-2 RNA

DMHBI + !control RNA

Population levelOur Biobricks:BBa_K1368010! BBa_K1368011

Single cell level

Improvements in I/O modules

Elegant designHighlights

• Carefully arranging the order of parts

• Reusing the existing parts and pathways

Balance of biology and engineeringBiological modules:overlapping and closely related

Module A

Module BM

odule C

Module A

Module B

Module C

Engineering modules: independent

Highlights

Balance of biology and engineering

Module A

Module BM

odule C

time

• Simplified like the biological modules

• Independent like the engineering modules

Highlights

Synthetic biological system with rewirable circuits is like Transformer!

Highlights

—Other implications of the Rewirable Circuits

Giving circuit adaptability is like giving it intelligence. What if it suddenly begins making variations of its own?

Policy & Practices

Policy & PracticesWhose Intellectual Property Right?

•Circuit designer

•Circuit user

•…Nobody?

A model providing stimulus for thinking

Awareness

PositiveNegative

StagePositive

Negative

2nd-dimension 3rd-dimension 4th-dimension

A 4-Dimensional model to help iGEMers think:

Policy & Practices

OutreachPolicy & Practices

Achievements

• Our input module, processing module and output module worked as we expected.

• We documented and submitted 13 new standard parts.

• We improved the characterization of 3 existing BioBrick Parts.

• Our team and HUST-China helped each other in lab work.

• We held a meetup this August in which 15 teams took part.

• We proposed a concept of rewirable circuit which can process different information with one module.

Acknowledgement

Instructors:Sponsors and departments:

Advisors:

AcknowledgementOur Team!

Thanks