Electrochemical sensor using intercalative, redox-active moieties

A method of detecting one or more single-base sequence lesions in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded, derivatized nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded derivatized nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded, derivatized nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded, derivatized nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded, derivatized nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in the target sequence.

A method of detecting one or more single-base sequence lesions in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety, wherein the intercalative, redox-active moiety is intercalatively stacked and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in the target sequence.

A method of detecting one or more single-base mismatches in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded, derivatized nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded derivatized nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence mismatches is indicative of the presence or absence of one or more single-base sequence mismatches in the target sequence.

A method of detecting one or more single-base sequence lesions in a portion of a gene sequence, wherein the one or more single-base sequence lesions is associated with a disease, comprising:

• a) contacting a first single stranded nucleic acid probe sequence, comprising a derivatized portion of the sequence of a gene, with a second single stranded nucleic acid target sequence comprising a wild type portion of the gene, to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in the gene sequence wherein the base stacking perturbation or single-base sequence lesion is associated with a disease.

A method of detecting one or more naturally occurring DNA lesions in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded nucleic acid probe sequence containing a DNA lesion with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more DNA lesions is indicative of the presence or absence of one or more DNA lesions in the target sequence.

A method of detecting one or more single-base sequence lesions in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an addressable multielectrode array, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the addressable multielectrode array, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the addressable multielectrode array, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in the target sequence.

A method of detecting one or more single-base sequence lesions in a target nucleic acid sequence, comprising:

• a) contacting a first single stranded nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an ultramicroelectrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the ultramicroelectrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the ultramicroelectrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in the target sequence.

A method of detecting one or more single-base sequence lesions in a hybrid of DNA and RNA, comprising:

• a) contacting a first single stranded nucleic acid probe sequence with a second single stranded nucleic acid target sequence to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of the one or more single-base sequence lesions in a hybrid of DNA and RNA.

A method of detecting one or more single-base sequence lesions in a portion of a derivatized wild type target nucleic acid sample, comprising:

• a) contacting a first single stranded nucleic acid probe sequence with a second single stranded nucleic acid target sequence, comprising a portion of a derivatized wild type target nucleic acid sample, to form a duplex, wherein the second single stranded nucleic acid target sequence is hybridized to a monolayer of first single stranded nucleic acid probe sequences on an electrode, wherein the monolayer is prepared by first attaching a duplex comprising the first single stranded nucleic acid probe sequence to the electrode, and dehybridizing the duplex, such that the first single stranded nucleic acid probe sequence remains attached to the electrode, and wherein the hybrid of the first single stranded nucleic acid probe sequence and the second single stranded target sequence form a double stranded nucleic acid-modified film;
• b) immersing the nucleic acid-modified film in a solution comprising an intercalative, redox-active moiety and a non-intercalative redox-active moiety;
• and
• c) measuring the electrical current or charges of the catalytic reduction of the non-intercalative, redox-active species by the intercalative, redox-active moiety wherein a difference of the electric current or charge relative to the electrical current or charge of a nucleic acid duplex not having one or more single-base sequence lesions is indicative of the presence or absence of one or more single-base sequence lesions in a portion of the derivatized wild type target nucleic acid sample.