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Dresden University of Technology

• T. Hinze

Computing with Molecules at Dresden University of Technology

Thomas Hinze

Dresden University of Technology Computer Science Department Theoretical Computer Science email: hinze@tcs.inf.tu-dresden.de www.molecular-computing.de

1/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Outline

Who we are

• part of Biosaxony Network in Dresden
• research group of 10 persons
• group leader: Dr. Monika Sturm

What we do

aims and visions

projects and results

scientific collaborations

teaching activities

BioInnovation Centre

www.biosaxony.com

Genetics Nanotechnology Bioinformatics Artificial Intelligence Theoretical Computer Science foundations

• approx. 80 companies

external partners Max Planck Institute of Molecular Cell Biology and Genetics

2/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Research Group Computing with Molecules

Aims and Visions

• modelling and simulation of molecular biological processes
• investigation, description, and optimization of models for computation
• biomolecule based algorithmic design

bridging gap between theoretical models and lab implementations Projects and Results

• wetware solution of the knapsack problem
• simulation system for phenomena undergoing side effects (Sisyphus)
• simple artificial chemistry experimental system (Saces)
• draft of universal programmable DNA based computer (TT6)
• library of data parallel algorithms based on Chomsky grammars
• genetic computing via microbial circuits in vivo

3/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Knapsack Problem

NP-complete, exponential need of resources, combinatorics Problem Definition There are

• natural numbers

and reference number

Is there a subset

• ✎

with

? Explanation

: weights of objects

• .

Is there a possibility to pack a selection of these objects into the knapsack and to meet the overall weight

exactly? Example

.

9 9 5 7 7

2 2

a = 7 a = 2 b = 9

1 2 3

a = 5

?

• bject 2
• bject 1
• bject 3

"yes" solution 4/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Wetware Solution of the Knapsack Problem

Brute Force Approach

• encode

into DNA double strands by length (

• ✁

)

• generate all solution candidates by a controlled

split-and-combine strategy

• separate the final DNA pool by length
• detect DNA at Starter length

• ✁

and answer yes

problem instance of size

• ✓

implemented in vitro

exponential need of resources moves from time to space

limited scalability of the algorithm because of side effects and amount of DNA

by length separation

10 20 30 5 15 25 35

a1 Starter Starter a1 Starter Starter Starter Starter Starter a1 a1 a2 a2 a2 Starter Starter Starter Starter Starter Starter Starter Starter a1 a2 a1 a2 a1 a2 a1 a2 DNA operations DNA operations DNA operations a3 a3 a3 a3 a3

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Dresden University of Technology

• T. Hinze

Modelling DNA Molecules

Primary Structure

• word over
• ✁

Secondary Structure

• set of nucleotides

• ✁

• relation of covalent bonds

• relation of hydrogen bonds

• inductive definition of

and

mathematical model to describe term (graph) rewriting rules

self assembly towards linear and nonlinear polymers

allows formalization of DNA operations on a moderate abstraction level

nucleotides without hydrogen bonds nucleotides with hydrogen bonds

P H H P

3’ 5’ 5’ 3’

A G G T C C G T A T A T C T A P H A T A G G T C C G T A T C T A P H T T A G A C T G C A A G T A C G T T G C A T G G A C C T H P T T A G A C T G C A A G T A C G T T G C A T G G A C C T H P

covalent bonds hydrogen bonds T C T A A T A G C T C G G T C C A G G T A C G C G T A A T T A T G T C G G T C A A A T

6/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

DNA Operations (I)

Generating DNA Strands

• synthesis (oligonucleotides)

5’−ACGGAAC−3’ A C G G A A C

• isolation . . . . . . . . . . . . . . . . . . . .

. . . . . . . . . . . .from organisms Merging and Aliquoting

• union/split

. . . . . . . . . . . . . . . . . . Modifying Hydrogen Bonds

• annealing (hybridization)

. .

T T G C C T T A C G G A A C T A T C G G C G C A T A T C

• melting (denaturation)

. . . .

T A T C G G C G C A T A T C T T G C C T T A C G G A A C

7/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

DNA Operations (II)

Enzymatic Reactions

• ligation (concatenation)

. . . . . . . . . .

T A C G G C A

P

T A T T A

P

T A C G G C T A A T T A

• digestion (cleavage)

. . . . . . . . . . . . . . .

A T G C C G G C C G A T A T G C G C

P

C G C G A T

P

HinP1 I

G C C G G C C G

• labeling (strand end modification)

A T G C C G G C C G A T A T G C C G G C C G A T

P P

+ 5’−phosphate

• polymerisation (blunting)

. . . . . . . . . .

A G C G G C C A A T G C C G G C

• PCR (polymerase chain reaction) .

. . . . . . . . . duplicate strands Separating and Analyzing of DNA Pools

• affinity purification (sep. by biotin)

A C G C G T A A T G C T C

B T

A A T T A T C

B T

A A T T A + 5’−biotin

• gel electrophoresis (sep. by length)

. . .sort and detect strands

• sequencing (read out strand)

. . . . .

A C G G A A C 5’−ACGGAAC−3’

8/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Side Effects of DNA Operations

(min. # copies) artifacts (diff. from structure) loss of linear DNA strands by forming hairpins, insertion deletion (% deletion rate, max. length of deletion) point mutation (% mutation rate) sequence) in DNA (differences mutations classification of side effects synthesis

• perations performed with

state of the art laboratory techniques bulges, loops, junctions, and compositions of them (% loss rate of tube contents)

• lin. DNA

(differences from incomplete reaction (% unprocessed strands) failures in reaction procedure unspecificity (% error rate, maximum difference) supercoils strand instabilities caused by temperature or pH impurities by rests of reagences loss of DNA strands (% loss rate of tube contents) perfect specification

• f reaction)

undetectable low DNA concentration in brackets: statistical parameters : supported in simulation tool : significant side effect caused by the operation annealing melting union ligation digestion labeling polymerisation PCR gel electrophoresis affinity purification

9/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

A PCR Example (I)

correct synthesized strands incorrect synthesized strands carrying point mutations and deletions 941 copies of template2) 59 strands of template2) 91 strands of primer1, (totally 90 strands of primer2, 942 copies of template1,

5

7909 copies of primer1,

5 6 7 941

(7910 copies of primer2,

942

58 strands of template1,

7910 7909

1000 copies template1 Synthesis Synthesis Synthesis 1000 copies template2 Synthesis Union Union Union 8000 copies primer1 8000 copies primer2

10/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

A PCR Example (II)

3rd PCR cycle 1st PCR cycle 2nd PCR cycle lane 1: PCR negative control lanes 2, 3, 4: PCR product (simulation screenshot) lane 5: 50bp marker 100 50 4 3 2 1 short double stranded PCR fragments) strands with deletions resulting in too unwanted strands (including amplified primer rests template (5965 copies) correct amplified double stranded 5

1 1 2 2 6 2713 2739 5965

• min. bonding rate: 50%
• max. length: 100bp

Melting Polymerisation Annealing

• min. bonding rate: 50%

Melting Polymerisation

• max. length: 100bp

Annealing

• min. bonding rate: 50%
• max. length: 100bp

Annealing Polymerisation Melting

11/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Simple Artificial Chemistry Experimental System

Idea

• behaviour of ideal gases can compute

Principle

• set of particles

molecules with parameters (

• ✂✂✁

b

)

• randomly placed/speeded due to

Maxwell-Boltzmann distribution

• set of reactions and global parameters

activation

(

• r

can be empty)

• algorithm: Brownian movement (random walk) with

elastic/inelastic collisions, retains momentum and energy

• analysis: animation, log, report, histogram

suitable for solution of NP-complete problems

info and software download: http://saces.yce.ch

x y z

12/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

cleavage with restriction enzymes restriction site: and ligation

1

α x1 β1x2 α2

1

y

1

α x1 β1x2 α2

1

y

1

α x1 β1x2 α2

1

y

1

α x1 β1x2 α2

1

y

1

α x1 α2

1

y β1x2

1

α x1 β1x2 α2

1

y β2y2 β2y2 β2y2 β2y2 β2y2 β2y2 α1 β1 α2 β2

Splicing Operation

• DNA recombination by cleavage and ligation can compute
• enables nondeterministic computation based on term rewriting
• main operation of programmable splicing systems (EH systems)

Definition (T. Head, 1987) Let

• an alphabet and

,

two symbols

• . A splicing rule
• ver
• is a word

with

,

. We define for each

and for words

• ✟

:

iff

13/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Ti splicing

• peration

filtering strands for , j = i Ai Ri Fj strands from distribution T , j = i

j

iterated in Tj , j = i , i = 1, ..., 5 initialization

Universal Distributed Splicing System (TT6)

Properties

• computational complete
• finite system components
• static structure
• programmable by Chomsky grammars
• massive data parallel processing
• use of linear DNA strands
• computational results provided

in separate test tube

• ✁
• system can be described using

well-known DNA operations

• minimization of distributed strands
• PCR based filtering method

14/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Principle of Univers. Distributed Splicing System TT6

Filter 4 Filter 3 Filter 2 Filter 1 Filter 6 Filter 5 Filter 5 Filter 4 Filter 3 Filter 2 Filter 1 Filter 6 Filter 5 Filter 3 Filter 2 Filter 1 Filter 4 Filter 6 Filter 5 Filter 4 Filter 3 Filter 2 Filter 1 Filter 6 Filter 5 Filter 4 Filter 3 Filter 2 Filter 1

: Platzhalter für spezifische Gruppen von DNA-Sequenzen

Filter 6

Bioreaktor 1 Bioreaktor 3 Bioreaktor 4 Bioreaktor 5 Bioreaktor 6 Bioreaktor 2

15/17 Computing with Molecules at TUD

Dresden University of Technology

• T. Hinze

Genetic Computing via Microbial Circuits in vivo

• construct connection free logic gates (NOT, NAND, FF

,

) using cell signalling pathways and controlled gene expression Example NOT Gate

• rganism vibrio fischeri, signalling network
• f promoter (
• ) and repressor (

) proteins

• input: signalling molecules AHL

(N-acryl homoserine lactones)

• utput: gfp (green fluorescence protein)

AHL 1 1 gfp AHL gfp t M cell i cell j signal sensor

• utput

regulatory circuit

pTSM b2 pCIRb PL* Ptrc PL* PL* Plux Lux I pAHLb AHL AHL AHL lux I lux R lac I lac I cl857 gfp AHL cellular extra

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Dresden University of Technology

• T. Hinze

Commitment outside Biosaxony

Scientific Collaborations

• European Molecular Computing Consortium (EMCC)
• Leiden Institute of Advanced Computer Science (LIACS)
• Vienna University of Technology, Theory and Logic Group
• Berne University of Applied Sciences, School of Engineering
• Fraunhofer Institute for Integrated Circuits (IIS)
• Philipps University Marburg, Dpt. Clinical Cytobiology and Cytopathology

Teaching Activities

• Th. Hinze, M. Sturm.

Rechnen mit DNA – Eine Einf¨ uhrung in Theorie und Praxis. Oldenbourg Wissenschaftsverlag M¨ unchen, 2004

The German-speaking book provides a comprehensive and syste- matic introduction into the interdisciplinary field of DNA computing including its mathematical and molecular biological background. The transfer of basic knowledge about DNA computing is com- pleted by the detailled introduction of models, methods, and techniques that prepare implementations in vitro. Particularly process simulations on a submolecular level are considered.

17/17 Computing with Molecules at TUD