Patent EP0410618A2 successfully refreshed from Google Patents.
EP0410618A2
Expired / Lapsed
EP
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Method for visualizing the base sequence of nucleic acid polymers
Assignee: Research Foundation of City University of New York
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Filing: 1990-07-17
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Granted: —
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Expires: 2010-07-17
Abstract
Abstract A method for determining and visualizing the base sequence of nucleic acid polymers, especially DNA and RNA, with a scanning probe microscope having a tip, the method including replacing the oxygen in the nucleic acid polymer with sulfur, complexing the sulfur with a metal such as mercury, and passing the tip over the complexed polymer in a scanning path to measure the potential and record the difference in electrical conductivity at preselected increments along the scanning path.
Inventors
- 👤 Stuart M. Lindsay
- 👤 Manfred Philipp
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-28 14:15
CPC Classifications
G; G01; G01Q; G01Q60/00; G01Q60/10; C; C07; C07H; C07H19/00; C07H19/02; C07H19/04; C07H19/06; C07H19/10; C07H19/16; C07H19/20; C07H23/00; Y; Y10; Y10S; Y10S435/00; Y10S435/81; Y10S977/00; Y10S977/84; Y10S977/849; Y10S977/852; Y10S977/853
Claims Analysis
Total Claims: 26
Claim 1
Independent Claim
54 words
A method of visualizing the base sequence of nucleic acid polymers comprising obtaining the nucleic acid polymer for sequencing, preparing a derivative thereof which has a reduced oxygen content, contacting the derivative with a metal or metal compound to form a metal-complexed nucleic acid polymer and visualizing the polymer under a scanning probe microscope.
Claim 2
Independent Claim
54 words
A method of visualizing the base sequence of nucleic acid polymers comprising obtaining the nucleic acid polymer for sequencing, preparing a derivative thereof which has a reduced oxygen content, contacting the derivative with a metal or metal compound to form a metal-complexed nucleic acid polymer and visualizing the polymer under a scanning probe microscope.
Claim 3
Independent Claim
159 words
A method for the determination of the nucleotide sequence of DNA and RNA with a scanning probe microscope having a tip, said method comprising: (a) contacting a DNA or RNA sample with a polymerase in the presence of a sulfur substituted varient of a nucleoside triphosphate in said DNA or RNA sample to form an amplified DNA or RNA polymer containing a sulfur containing nucleotide; (b) contacting the amplified polymer of step (a) with a soluble organometallic compound to form a metal-complexed nucleic acid polymer; (c) disposing said metal-complexed nucleic acid polymer on a substrate which is atomically flat within micron distances; (d) passing said tip of said scanning microscope in a probe path over the surface of said polymer to measure electrical potential therebetween; and (e) recording the difference in electrical potential so measured at preselected increments along said scanning probe path to identify the location of nucleoside bases to determine the sequence of said DNA nucleoside bases.
Claim 4
Independent Claim
159 words
A method for the determination of the nucleotide sequence of DNA and RNA with a scanning probe microscope having a tip, said method comprising: (a) contacting a DNA or RNA sample with a polymerase in the presence of a sulfur substituted varient of a nucleoside triphosphate in said DNA or RNA sample to form an amplified DNA or RNA polymer containing a sulfur containing nucleotide; (b) contacting the amplified polymer of step (a) with a soluble organometallic compound to form a metal-complexed nucleic acid polymer; (c) disposing said metal-complexed nucleic acid polymer on a substrate which is atomically flat within micron distances; (d) passing said tip of said scanning microscope in a probe path over the surface of said polymer to measure electrical potential therebetween; and (e) recording the difference in electrical potential so measured at preselected increments along said scanning probe path to identify the location of nucleoside bases to determine the sequence of said DNA nucleoside bases.
Claim 5
Dependent Claim
37 words
A method according to claim 2 in which said organometallic compound contains a metal selected from the group consisting of mercury, silver, bismuth, cadmium, cerrium, cobalt, copper, iron, lanthanum, manganese, nickle, lead, platinum, tin, titanium, and zinc.
Claim 6
Dependent Claim
13 words
A method according to claim 2 in which said microscope is a STM.
Claim 7
Dependent Claim
12 words
A method according to claim 2 in which said microscope is AFM.
Claim 8
Independent Claim
111 words
A method for visualizing the base sequence of nucleic acid polymers with a scanning probe microscope having a tip, said method comprising replacing the oxygen in a nucleic acid polymer with sulfur to form a sulfur-containing polynucleotide, placing said sulfur-containing polynucleotide into a solution containing a soluble mercury compound to complex said mercury-containing compound with the sulfur-containing polynucleotide; disposing said mercury-complexed nucleic acid polymer on a substrate which is atomically flat within micron distances; passing the tip of said scanning microscope in a path over the surface of said polymer to measure potential therebetween; and recording the difference in electrical conductivity so measured at preselected increments along said scanning probe path.
Claim 9
Independent Claim
111 words
A method for visualizing the base sequence of nucleic acid polymers with a scanning probe microscope having a tip, said method comprising replacing the oxygen in a nucleic acid polymer with sulfur to form a sulfur-containing polynucleotide, placing said sulfur-containing polynucleotide into a solution containing a soluble mercury compound to complex said mercury-containing compound with the sulfur-containing polynucleotide; disposing said mercury-complexed nucleic acid polymer on a substrate which is atomically flat within micron distances; passing the tip of said scanning microscope in a path over the surface of said polymer to measure potential therebetween; and recording the difference in electrical conductivity so measured at preselected increments along said scanning probe path.
Claim 10
Dependent Claim
22 words
A method according to claim 6 in which said nucleic acid polymer is selected from the group consisting of DNA and RNA.
Claim 11
Dependent Claim
13 words
A method according to claim 7 in which said microscope is a STM.
Claim 12
Dependent Claim
14 words
9 . A method according to claim 7 in which said microscope is AFM.
Claim 13
Dependent Claim
30 words
A method according to claim 6 in which said substrate is prepared by depositing Angstrom amounts of gold epitaxially on freshly cleaned scratch-free mica in an ultra high vacuum system.
Claim 14
Dependent Claim
34 words
A method according to claim 10 in which said gold-bearing mica is preheated to about 300°C in a oil-free system at least 10⁻⁹ torr, removed from said system, and stored under argon until needed.
Claim 15
Dependent Claim
15 words
A method according to claim 6 in which said tip is formed of Pt-Ir wire.
Claim 16
Dependent Claim
94 words
A method according to claim 12 in which said tip is pretreated by immersing said tip into a solution of 3 M NaCN and 1 M NaOH, applying 20 volts of a c to said immersed tip until 0.5 Amperes rms. flows therethrough; stopping the current flow; applying small bursts of current to said tip until the lower region thereof parts; pressing said tip through a film of hard wax preheated to a temperature of from 100°C to about 200°C to deposit wax on said tip; and removing said wax-coated tip from said wax.
Claim 17
Independent Claim
142 words
A method for visualizing the base sequence of a nucleic acid polymer selected from the group consisting of DNA and RNA with a scanning tunneling microscope having a tip operatively associated therewith, said method comprising placing said nucleic acid polymer in a solution containing a compound having an atom capable of replacing the oxygen atom in said polymer; dissolving said oxygen-reduced polymer into a buffer solution; depositing said polymer-containing buffer solution in a cell having a substrate: introducing sodium p-hydroxymercuribenzoate into said cell; inserting a Pt-wire counter electrode into said cell under a 2V bias to cause hydrogen to evolve therefrom; lowering said tip toward said substrate to measure the difference in electrical conductivity at preselected increments thereof along a predefined path; and translating said differences in electrical conductivity into a visual pattern representing the base sequence of said nucleic acid polymer.
Claim 18
Independent Claim
142 words
A method for visualizing the base sequence of a nucleic acid polymer selected from the group consisting of DNA and RNA with a scanning tunneling microscope having a tip operatively associated therewith, said method comprising placing said nucleic acid polymer in a solution containing a compound having an atom capable of replacing the oxygen atom in said polymer; dissolving said oxygen-reduced polymer into a buffer solution; depositing said polymer-containing buffer solution in a cell having a substrate: introducing sodium p-hydroxymercuribenzoate into said cell; inserting a Pt-wire counter electrode into said cell under a 2V bias to cause hydrogen to evolve therefrom; lowering said tip toward said substrate to measure the difference in electrical conductivity at preselected increments thereof along a predefined path; and translating said differences in electrical conductivity into a visual pattern representing the base sequence of said nucleic acid polymer.
Claim 19
Dependent Claim
15 words
15 . A method according to claim 14 in which said compound is a thiophosphate.
Claim 20
Dependent Claim
13 words
A method according to claim 15 in which said thiophosphate is alpha-thio-deoxynucleoside triphosphate.
Claim 21
Independent Claim
130 words
A method for visualizing the base sequence of nucleic acid polymers with a scanning probe microscope having a tip, the method comprising: replacing the oxygen in a nucleic acid polymer to form a sulfur-containing polynucleotide; placing said sulfur-containing polynucleotide into a solution containing a soluble cation-containing compound, said cation being selected from the group consisting of mercury, silver, bismuth, cadmium, cerrium, cobalt, copper, iron, lanthanum, manganese, nickle, lead, platinum, tin, titanium and zinc, to complex said cation-containing compound with said sulfur-containing polynucleotide; disposing said cation-complexed nucleic acid polymer on an atomically flat substrate; passing the tip of said scanning microscope in a path over the surface of said polymer to measure potential therebetween; and recording the difference in electrical conductivity so measured at preselected increments along said scanning probe path.
Claim 22
Independent Claim
130 words
A method for visualizing the base sequence of nucleic acid polymers with a scanning probe microscope having a tip, the method comprising: replacing the oxygen in a nucleic acid polymer to form a sulfur-containing polynucleotide; placing said sulfur-containing polynucleotide into a solution containing a soluble cation-containing compound, said cation being selected from the group consisting of mercury, silver, bismuth, cadmium, cerrium, cobalt, copper, iron, lanthanum, manganese, nickle, lead, platinum, tin, titanium and zinc, to complex said cation-containing compound with said sulfur-containing polynucleotide; disposing said cation-complexed nucleic acid polymer on an atomically flat substrate; passing the tip of said scanning microscope in a path over the surface of said polymer to measure potential therebetween; and recording the difference in electrical conductivity so measured at preselected increments along said scanning probe path.
Claim 23
Dependent Claim
12 words
A method according to claim 15 in which said cation is mercury.
Claim 24
Dependent Claim
17 words
A method according to claim 18 in which said mercury is contained in a soluble mercury salt.
Claim 25
Dependent Claim
18 words
A method according to claim 18 in which said mercury is contained in a soluble alkyl mercury compound.
Claim 26
Dependent Claim
18 words
A method according to claim 18 in which said mercury is contained in a soluble aryl mercury compound.
References Cited (Backward Citations)
No citation records indexed.
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