WO2003073096A1 - Portes logiques et reseaux moleculaires a base d'oligonucleotide - Google Patents

Portes logiques et reseaux moleculaires a base d'oligonucleotide Download PDF

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Publication number
WO2003073096A1
WO2003073096A1 PCT/US2003/005506 US0305506W WO03073096A1 WO 2003073096 A1 WO2003073096 A1 WO 2003073096A1 US 0305506 W US0305506 W US 0305506W WO 03073096 A1 WO03073096 A1 WO 03073096A1
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Prior art keywords
gate
logic
loop
logical
stem
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PCT/US2003/005506
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English (en)
Inventor
Milan N. Stojanovic
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The Trustees Of Columbia University In The City Of New York
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Priority to AU2003213250A priority Critical patent/AU2003213250A1/en
Publication of WO2003073096A1 publication Critical patent/WO2003073096A1/fr

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Classifications

    • H—ELECTRICITY
    • H03—ELECTRONIC CIRCUITRY
    • H03K—PULSE TECHNIQUE
    • H03K19/00—Logic circuits, i.e. having at least two inputs acting on one output; Inverting circuits
    • H03K19/0005—Modifications of input or output impedance
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813—Hybridisation assays

Definitions

  • the present invention provides an oligonucleotide logic gate.
  • General allosteric control of deoxyribozymes (DNA-based catalysts 14 ), with phosphodiesterase activity by oligonucleotides is important in this context 15 , because the product oligonucleotide (output) of one catalyst could be used as an allosteric effector (input) of another catalyst, thereby allowing communication between various elements of the multifunctional platform without a change in phase.
  • a logic gate comprising at least one input, at least one output, at least one oligonucleotide with catalytic activity and at least one stem-loop which controls the catalytic activity of the gate, wherein each said output is capable of at least two different output states, said states depending on the catalytic activity of the gate.
  • the logic gate may be arranged and used to detect a disease marker, wherein the disease marker has been translated into an oligonucleotide.
  • the logic gate may be arranged and used to signal a disease marker, wherein the disease marker has been translated into an oligonucleotide.
  • a plurality of logic gates of the type described above is provided, wherein the output of one gate is arranged as the input of another gate.
  • the product of one gate may be arranged to be the input of another gate.
  • a method of performing a logical operation is provided using a logic gate having catalytic activity, at least one input, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • a method of performing a logical operation is provided using a logic gate having catalytic activity, at least one input, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:-
  • Figure 1 Basic concept: input oligonucleotides IA and IB result in the presence or absence of output fluorescent product OF depending on the interactions with deoxyribozyme-based logic gates.
  • Figure 2 Fluorogenic cleavage of double end-labeled substrate by deoxyribozymes 12E or 8-17 into products OF and OR. Fluorescein (F) emission is quenched by distance dependent fluorescence resonance energy transfer to tetramethylrhodamine (R), and upon cleavage fluorescence increases (larger font F).
  • Fluorescein (F) emission is quenched by distance dependent fluorescence resonance energy transfer to tetramethylrhodamine (R), and upon cleavage fluorescence increases (larger font F).
  • Figure 2A Deoxyribozyme Logic: Basic Technologies.
  • Figure 3 Single input sensor gate (A) is activated by the input oligonucleotide IA.
  • IA input oligonucleotide
  • Insert in box schematically represents inactive gate with closed loop (output 0) and active gate with open loop (output 1);
  • Figure 4 Single-input NOT gate ( -, B ) is constructed through substitution of a non- conserved loop in the deoxyribozyme with beacon stem loop complementary to the input.
  • Deoxyribozyme is inactive in the complex with In, while 1 A has only minimal inhibitory influence; insert in box schematically represents active gate with closed loop (output 1) and inactive gate with open loop (output 0);
  • Figure 4A Deoxyribozyme - Based Sensors for Proteins: NOT Streptavidine Gate.
  • Figure 4B OR Gate and NANO Gate.
  • FIG. 5 AND gate (A «B) is constructed through attachment of two loops complementary to input oligonucleotides to the 5' and 3' ends of the deoxyribozyme; deoxyribozyme is active only if both inputs are present; insert in box schematically presents inactive gate (output 0) with either one or both loops closed, and active gate with both loops open (output 1); Graph shows fluorescence spectra (relative intensity vs.
  • Figure 6 (a) A sensor-inhibitor AND NOT gate (A* ⁇ B) is constructed through attachment of two loops complementary to input oligonucleotides, one at the 5' end, one at the non-conserved loop; catalytic activity in solution is present only if I A is present and I
  • Figure 7 XOR gate (AvB) as a combination of A -'B and B*- with same inputs and output; catalytic activity is present in solution if either IA or IB is present, but not both; insert in box schematically represents the two active states (output 1) of the XOR system, when only one oligonucleotide is present (second or third), and two inactive states (output 0) with either neither (first) or both (fourth) oligonucleotides present; Graph shows fluorescence spectra (relative intensity vs.
  • Figure 8 Deoxyribozyme-based half-adder, demonstrating parallel operation of two logic gates; XOR gate will be active and yield product if only one of the inputs is present, not both; AND gate will be active only if both inputs are present. Substrates are engineered to allow for multicolor detection. (BH-black hole quenchers; F- fluorescein, R : rhodamine). Inserts represent corresponding truth table.
  • Figure 10 Operation of NOT streptavidine sensor gate and NOT oligonucleotide sensor gate in series (truth table with O F as an output). Substrate of upstream NOT streptavidine gate inhibits NOT oligonucleotide gate, while products do not. Note the translation of the output.
  • Figure 11 Construction of a system that behaves and NAND gate from: 1. Clocking Module, which synchronizes activity of NOT oligonucleotide-sensor gate with AND Gate; 2. AND gate, that cleaves substrate constrained into stem-loop structure, and yields product which inhibits NOT oligonucleotide gate; 3. NOT oligonucleotide sensor gate, which is inhibited by two oligonucleotides, one is the substrate of clocking deoxyribozyme, and the other product of AND gate. Note that NOT module is inactive only if AND module is active, i.e. when both input oligonucleotides are present in the solution. Insert in NOT module describes truth table of a NOT gate in this network, behaving as a NAND gate in regard to input E, S and I denote enzymes, substrates and inputs of corresponding modules.
  • Figure 12 Basic principle of deoxyribozyme chain reaction; Input oligonucleotide activates sensor gate, which cleaves substrate into another input oligonucleotide and an output oligonucleotide for downstream gate. Thus, each input oligonucleotide starts a chain reaction, as sensor gates have multiple turnovers.
  • Figure 14 A network of three AND gates, with outputs of two AND gates connected as inputs to a third AND gate.
  • a logic gate comprising at least one input, at least one output, at least one oligonucleotide with catalytic activity and at least one stem-loop which controls the catalytic activity of the gate, wherein each said output is capable of at least two different output states, said states depending on the catalytic activity of the gate.
  • the configuration of the stem-loop preferably determines the output state.
  • the gate preferably has one input, and a first output state when the stem-loop is closed and a second output state when the stem-loop is open.
  • the first output state may correspond to a logical off and the second output state may correspond to a logical on.
  • the first output state may correspond to a logical on and the second output state may correspond to a logical off.
  • the output of the gate may comprise a fluorescent readout, electromagnetic readout, colorimetric readout, radiation readout, a light emission readout, and/or an ultraviolet spectral change readout.
  • the output of the gate may comprise a material whose conductivity changes to indicate the output states.
  • the output of the gate may comprise a material whose magnetization changes to indicate the output state.
  • the stem-loop may comprise an oligonucleotide.
  • the oligonucleotide may comprise a peptide nucleic acid.
  • the logic gate may comprise peptide nucleic acid.
  • the stem- loop may comprise peptide nucleic acid.
  • the logic gate may comprise DNA.
  • the logic gate may comprise RNA.
  • the DNA may comprise natural DNA.
  • the DNA may comprise synthetic DNA.
  • the RNA may comprise natural RNA.
  • the RNA may comprise synthetic RNA.
  • the logic gate may comprise both natural and synthetic nucleotides.
  • At least one input may comprise an oligonucleotide.
  • the logic gate may further comprise at least one input based on hybridization.
  • the logic gate may further comprise at least one input based on complementary base pair formation.
  • At least one output may comprise an oligonucleotide.
  • the number of inputs may be at least two.
  • the gate may be a logical AND gate, comprising at least two inputs, and being in a logical on state only if all inputs are in the same one of two states.
  • the gate may be a logical AND NOT gate, comprising two inputs, and being in a logical on state if and only if one input is in a certain one of two states.
  • the logic gate may have one input, and form a logical NOT gate, being in a logical on state if the input is in a certain one of two states.
  • the logic gate may comprise more than two inputs, wherein the gate is in a logical on state if at least one constituent stem-loop is in an open or closed state.
  • the logic gate may comprise a substrate binding region, wherein substrate binding is inhibited when the stem-loop is in the closed state.
  • the substrate binding may be inhibited when the stem-loop is in the open state.
  • the gate may be a logical sensor gate, wherein an input is transduced into an output.
  • the logic gate may have a catalytic core region, wherein the stem-loop is attached to the catalytic region of the gate.
  • the gate may be a logical NOT gate.
  • the logic gate may be arranged and used to detect a disease marker, wherein the disease marker has been translated into an oligonucleotide.
  • the logic gate may be arranged and used to signal a disease marker, wherein the disease marker has been translated into an oligonucleotide.
  • a plurality of logic gates of the type described above is provided, wherein the output of one gate is arranged as the input of another gate.
  • the product of one gate may be arranged to be the input of another gate.
  • a plurality of gates may have a common substrate.
  • the substrate of one gate may be the input of another gate.
  • a logic gate performing a catalytic function as a logic operation, said gate having at least one input and at least one output, said gate providing an output having a characteristic which depends on a characteristic of the input, said output characteristic being sufficient to be provided as an input characteristic to a second logic gate.
  • the gate may have at least two inputs.
  • the logic operation may be AND.
  • the logic operation may be XOR.
  • the logic operation may be a sensing operation and the gate may be a YES gate.
  • the gate may comprise a deoxyribozyme.
  • the gate may comprise a ribozyme.
  • the logic gate may comprise peptide nucleic acid.
  • the logic gate may comprise DNA.
  • the logic gate may comprise RNA.
  • the DNA may comprise natural DNA.
  • the DNA may comprise synthetic DNA.
  • the RNA may comprise natural RNA.
  • the RNA may comprise synthetic RNA.
  • the logic gate may comprise both natural and synthetic nucleotides.
  • the logic gate may further comprise a second logic gate, said second logic gate receiving as an input the output of the first logic gate.
  • a method of performing a logical operation is provided using a logic gate having catalytic activity, at least one input, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • a method of performing a logical operation is provided using a logic gate having catalytic activity, at least one input, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • a method of performing a logical AND operation is provided using a logic gate having catalytic activity, a plurality of inputs, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • a method of performing a logical AND operation is provided using a logic gate having catalytic activity, a plurality of inputs, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • a method of performing a logical AND NOT operation is provided using a logic gate having catalytic activity, a plurality of inputs, and an output capable of at least two different output states, said states depending on the catalytic activity of the gate, said logic gate further comprising at least one oligonucleotide and at least one stem-loop which controls the gate catalytic activity, which method comprises the steps of:
  • ligases are usually significantly larger enzymes (with one apparent exception) and therefore less practical.
  • Hamming distance in these oligonucleotide-based computation elements can be defined as number of mismatches that minimizes cross talk between two elements. Thus, at room temperature and high Mg 2+ concentrations, for 15- er oligonucleotides the Hamming distance can be realistically set at 3 for YES gates and 4 for NOT gates.

Abstract

La présente invention a trait à un ensemble de portes logiques à base de désoxyribozymes capables de générer n'importe quelle fonction booléenne. Les portes comportent des portes classiques NON et ET, et la porte OU exclusif plus complexe. Ces portes ont été construites grâce à une conception modulaire qui combine les tiges-boucles de balises moléculaires avec les désoxyribozymes de type marteau. Les portes comprennent des oligonucléotides en entrées et en sortie, permettant ainsi la communication entre différents éléments de calcul en solution. Le fonctionnement de ces portes est avantageusement relié à un affichage fluorescent.
PCT/US2003/005506 2002-02-21 2003-02-21 Portes logiques et reseaux moleculaires a base d'oligonucleotide WO2003073096A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU2003213250A AU2003213250A1 (en) 2002-02-21 2003-02-21 Oligonucleotide-based logic gates and molecular networks

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US3780302A 2002-02-21 2002-02-21
US10/037,803 2002-02-21

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Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5561071A (en) * 1989-07-24 1996-10-01 Hollenberg; Cornelis P. DNA and DNA technology for the construction of networks to be used in chip construction and chip production (DNA-chips)
US5955322A (en) * 1996-02-07 1999-09-21 Mount Sinai School Of Medicine Of The City University Of New York DNA-based computer

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5561071A (en) * 1989-07-24 1996-10-01 Hollenberg; Cornelis P. DNA and DNA technology for the construction of networks to be used in chip construction and chip production (DNA-chips)
US5955322A (en) * 1996-02-07 1999-09-21 Mount Sinai School Of Medicine Of The City University Of New York DNA-based computer

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
ADLEMAN: "Molecular computation of solutions to combinatorial problems", SCIENCE, vol. 266, 11 November 1994 (1994-11-11), pages 1021 - 1024, XP002043104 *
FITCH: "Calculating the expected frequencies of potential secondary structure in nucleic acids as a function of stem length, loop size, base composition and nearest-neighbor frequencies", NUCLEIC ACIDS RESEARCH, vol. 11, no. 13, 1983, pages 4655 - 4663, XP002963363 *

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