Existing “autonomous” system Existing “autonomous” system Sakamoto & Hagiya Sakamoto & Hagiya State transitions by molecules State transitions by molecules A transition table: A transition table: {S {S S’} S’} Starting from the initial state, calculate as many as possible Starting from the initial state, calculate as many as possible following states according to the transition table following states according to the transition table
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Existing “autonomous” system Sakamoto & Hagiya State transitions by molecules A transition table:{S S’} Starting from the initial state, calculate as.
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Existing “autonomous” systemExisting “autonomous” system
Sakamoto & HagiyaSakamoto & Hagiya
State transitions by moleculesState transitions by molecules
A transition table:A transition table: {S {S S’} S’}
Starting from the initial state, calculate as many as possibleStarting from the initial state, calculate as many as possiblefollowing states according to the transition tablefollowing states according to the transition table
Molecular implementationMolecular implementation
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
• Each state is a ss-DNA segmentEach state is a ss-DNA segment• Each transition S -> S’ is a sequence of S-complementaryEach transition S -> S’ is a sequence of S-complementaryand S’-complementary DNAand S’-complementary DNA• The transitions are performed in a PCR-like fashionThe transitions are performed in a PCR-like fashion
States are not physically separated from the symbols.States are not physically separated from the symbols.Subsequences of the alphabet codes represent different statesSubsequences of the alphabet codes represent different states
CTGGCTGG = = a combination of S1 and 0a combination of S1 and 0
States are not physically separated from the symbols.States are not physically separated from the symbols.Subsequences of the alphabet codes represent different statesSubsequences of the alphabet codes represent different states
GGCTGGCT = = a combination of S0 and 0a combination of S0 and 0
CTGGCTGG = = a combination of S1 and 0a combination of S1 and 0
States are not physically separated from the symbols.States are not physically separated from the symbols.Subsequences of the alphabet codes represent different statesSubsequences of the alphabet codes represent different states
States are not physically separated from the symbols.States are not physically separated from the symbols.Subsequences of the alphabet codes represent different statesSubsequences of the alphabet codes represent different states
1,10: 50 1,10: 50 bp ladder; 2: 101 input; 3: 010010 input; 4: S0-detector; 5: S1-detectorbp ladder; 2: 101 input; 3: 010010 input; 4: S0-detector; 5: S1-detector6: Computation result of 101 input; 7: Computation result of 010010 input6: Computation result of 101 input; 7: Computation result of 010010 input8: Computation result of 010100 input; 9: Computation result of 001000 input8: Computation result of 010100 input; 9: Computation result of 001000 input
150 150 bpbp
200 200 bpbp
S0-resultS0-result
S1-resultS1-result
InputInputdegradationdegradationproductsproducts
Reaction conditions: Reaction conditions: Environment: 120 Environment: 120 l of NEB4 buffer + 1 mM ATP, 18 l of NEB4 buffer + 1 mM ATP, 18 ooC, 80 minC, 80 minInput: 2.5 pmol; Detectors: 1.5 pmol each; Transition molecules: 20 pmol eachInput: 2.5 pmol; Detectors: 1.5 pmol each; Transition molecules: 20 pmol eachFok I: 12 units; T4 Ligase: 120 unitsFok I: 12 units; T4 Ligase: 120 units
S0-dS0-d
S1-dS1-d
Proof of Mechanism
A complete mixture:A complete mixture:Input (010100) ; S0-detector; S1-detector; T1,T2,T3,T4; Fok I; T4 DNA LigaseInput (010100) ; S0-detector; S1-detector; T1,T2,T3,T4; Fok I; T4 DNA Ligase
The gel shows a “component removal” experiment, where each component was The gel shows a “component removal” experiment, where each component was omitted from the complete mixture and the result was compared to the predicted omitted from the complete mixture and the result was compared to the predicted outcomeoutcome
1,12: 50 1,12: 50 bp ladderbp ladder2: complete mixture2: complete mixture3: No Input3: No Input4: No S0-detector4: No S0-detector5: No S1-detector5: No S1-detector6: No T16: No T17: No T27: No T28: No T38: No T39: No T49: No T410: No Fok I10: No Fok I11: No T4 DNA Ligase11: No T4 DNA Ligase
1 2 3 4 5 6 71 2 3 4 5 6 7Detectors are labeled with Detectors are labeled with 3232PP
S0-resultS0-result
S1-resultS1-result
S1-detectorS1-detector
S0-detectorS0-detector
There are possible “wrong” bands. Their origin is currently There are possible “wrong” bands. Their origin is currently being determined. At any rate, the correctness is >95%being determined. At any rate, the correctness is >95%
Exact error rate still needs to be determinedExact error rate still needs to be determined
Lanes:Lanes:1,7: 50 bp ladder1,7: 50 bp ladder2: 2: 3232P-S0-detectorP-S0-detector3: 3: 3232P-S1-detectorP-S1-detector4: Computation over 0100104: Computation over 0100105: Computation over 0101005: Computation over 0101006: Computation over 0010006: Computation over 001000