Device and method for analyzing streptavidin

Patent metadata

Field	Value
Publication number	US-9880161-B2
Application number	US-201314387431-A
Country	US
Kind code	B2
Filing date	Mar 21, 2013
Priority date	Mar 23, 2012
Publication date	Jan 30, 2018
Grant date	Jan 30, 2018

Abstract

The present invention provides a novel sensor for detecting streptavidin (SA). The nucleic acid sensor for analyzing SA of the present invention includes the following nucleic acid element that includes a catalyst nucleic acid molecule (D) that exerts a catalytic function and a binding nucleic acid molecule (A) that binds to SA. The nucleic acid element is a double-stranded nucleic acid element including a first strand and a second strand. The first strand (ss1) includes the binding nucleic acid molecule (A), a loop-forming sequence (L1), and the catalyst nucleic acid molecule (D) linked in this order. The second strand (ss2) includes a stem-forming sequence (S A ), a loop-forming sequence (L2), and a stem-forming sequence (S D ) linked in this order. In this nucleic acid element, in the absence of SA, the catalytic function of the catalyst nucleic acid molecule (D) is inhibited by stem formation in each of the stem-forming sequences (S A ) and (S D ), and in the presence of SA, the stem formation is released by a binding of the binding nucleic acid molecule (A) with the SA, and the catalytic function of the catalyst nucleic acid molecule (D) is exerted.

First claim

The invention claimed is:

1. A nucleic acid sensor for analyzing streptavidin, comprising
a nucleic acid element (II) that comprises a single-stranded nucleic acid element comprising a binding nucleic acid molecule (A) that binds to streptavidin, a loop-forming sequence (L1), a stem-forming sequence (S D ), a catalyst nucleic acid molecule (D) that exerts a redox function, a loop-forming sequence (L2), and a stem-forming sequence (S A ) linked in this order, wherein
a terminal region of the binding nucleic acid molecule (A) on the loop-forming sequence (L1) side is complementary to the stem-forming sequence (S A ),
a terminal region of the catalyst nucleic acid molecule (D) on the loop-forming sequence (L2) side is complementary to the stem-forming sequence (S D ),
the loop-forming sequence (L1) is non-complementary to the loop-forming sequence (L2), and

wherein, in the absence of streptavidin,
the redox function of the catalyst nucleic acid molecule (D) is inhibited by stem formation in each of the stem-forming sequences (S A ) and (S D ), and
in the presence of streptavidin,
the stem formation in each of the stem-forming sequences (S A ) and (S D ) is released by a binding of the streptavidin with the binding nucleic acid molecule (A), and the redox function of the catalyst nucleic acid molecule (D) is exerted,
a terminal region of the binding nucleic acid molecule (A) on the loop-forming sequence (L1) side and the stem-forming sequence (S A ) form a stem,
a terminal region of the catalyst nucleic acid molecule (D) on the loop-forming sequence (L2) side and the stem-forming sequence (S D ) form a stem, and
the loop-forming sequences (L1) and (L2) form an internal loop between the two stems.

2. The nucleic acid sensor according to claim 1 , wherein
the nucleic acid element (II) comprises, from the 3′ side thereof, the binding nucleic acid molecule (A), the loop-forming sequence (L1), the stem-forming sequence (S D ), the catalyst nucleic acid molecule (D), the loop-forming sequence (L2), and the stem-forming sequence (S A ) linked in this order,
a 5′ terminal region of the binding nucleic acid molecule (A) is complementary to the stem-forming sequence (S A ), and
a 5′ terminal region of the catalyst nucleic acid molecule (D) is complementary to the stem-forming sequence (S D ).

3. The nucleic acid sensor according to claim 1 , wherein the length of each of the loop-forming sequences (L1) and (L2) ranges from 1- to 30-nucleotides.

4. The nucleic acid sensor according to claim 2 , wherein
the length of the stem-forming sequence (S A ) ranges from 1- to 60-nucleotides, and
the length of the stem-forming sequence (S D ) ranges from 1- to 30-nucleotides.

5. The nucleic acid sensor according to claim 1 , wherein
the length of the binding nucleic acid molecule (A) ranges from 18- to 85-nucleotides.

6. The nucleic acid sensor according to claim 1 , wherein
the binding nucleic acid molecule (A) comprises the following polynucleotide (a1), (a2), (a3), or (a4):
(a1) a polynucleotide composed of a base sequence of any of SEQ ID NOs: 1 to 10,
(a2) a polynucleotide that is composed of a base sequence obtained by substitution, deletion, addition and/or insertion of at least one base in the base sequence of the polynucleotide (a1) and binds to streptavidin,
(a3) a polynucleotide that is composed of a base sequence with 50% or more identity with the base sequence of the polynucleotide (a1) and is bindable to streptavidin, and
(a4) a polynucleotide that is composed of a base sequence complementary to a base sequence that hybridizes to the base sequence of the polynucleotide (a1) under stringent conditions and is bindable to streptavidin.

7. The nucleic acid sensor according to claim 1 , wherein
the catalytic function of the catalyst nucleic acid molecule (D) is the catalytic function of an oxidation-reduction reaction.

8. The nucleic acid sensor according to claim 1 , wherein
the length of the catalyst nucleic acid molecule (D) ranges from 15- to 30-nucleotides.

9. The nucleic acid sensor according to claim 1 , wherein
the catalyst nucleic acid molecule (D) comprises the following polynucleotide (d1), (d2), (d3), or (d4):
(d1) a polynucleotide composed of a base sequence of any of SEQ ID NOs: 11 to 31 and 61 to 80,
(d2) a polynucleotide that is composed of a base sequence obtained by substitution, deletion, addition, and/or insertion of at least one base in the base sequence of the polynucleotide (d1) and exerts a catalytic function of an oxidation-reduction reaction,
(d3) a polynucleotide that is composed of a base sequence with 50% or more identity with the base sequence of the polynucleotide (d1) and exerts a catalytic function of an oxidation-reduction reaction, and
(d4) a polynucleotide that is composed of a base sequence complementary to a base sequence that hybridizes to the base sequence of the polynucleotide (d1) under stringent conditions and exerts a catalytic function of an oxidation-reduction reaction.

10. A device for analyzing streptavidin, comprising:
a base material;
a nucleic acid sensor; and
a detection part, wherein
the nucleic acid sensor and the detection part are disposed on the base material,
the nucleic acid sensor is the nucleic acid sensor according to claim 1 , and
the detection part is a detection part that detects the catalytic function of the catalyst nucleic acid molecule (D) in the nucleic acid sensor.

11. The device according to claim 10 , wherein the nucleic acid sensor is linked with the base material via a linker.

12. The device according to claim 10 , wherein the nucleic acid sensor is disposed on the detection part.

13. The device according to claim 10 , wherein the detection part detects a signal generated by the catalytic function of the catalyst nucleic acid molecule (D).

14. The device according to claim 13 , wherein the signal is an optical signal or an electrochemical signal.

15. The device according to claim 10 , further comprising:
a reagent part, wherein the reagent part comprises a substrate to the catalytic function of the catalyst nucleic acid molecule (D).

16. A method for analyzing streptavidin, comprising:
a contact step of causing a sample to be in contact with the nucleic acid sensor according to claim 1 ; and
a detection step of detecting the catalytic function of the catalyst nucleic acid molecule (D) in the nucleic acid sensor to detect streptavidin in the sample.

17. The method according to claim 16 , wherein the detection step is performed in the presence of a substrate to the catalytic function of the catalyst nucleic acid molecule (D).

18. A method for analyzing streptavidin, comprising:
a contact step of causing a sample to be in contact with the device according to claim 10 ; and
a detection step of detecting the catalytic function of the catalyst nucleic acid molecule (D) in the detection part of the device to detect streptavidin in the sample.

19. The method according to claim 18 , wherein the detection step is performed in the presence of a substrate to the catalytic function of the catalyst nucleic acid molecule (D).

Assignees

• Horii Katsunori
• Kaneko Naoto
• Akitomi Jou
• Kato Shintaro
• Waga Iwao
• Nec Solution Innovators Ltd

Inventors

• Horii Katsunori
• Kaneko Naoto
• Akitomi Jou
• Kato Shintaro
• Waga Iwao

Classifications

• C12Y111/01007

  Peroxidase (1.11.1.7), i.e. horseradish-peroxidase · CPC title

• G01N2333/36

  from Actinomyces; from Streptomyces (G) · CPC title

• G01N33/58

  involving labelled substances (G01N33/53 takes precedence) · CPC title

• C12N15/113

  Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)} · CPC title

• G01N33/54306 Primary

  Solid-phase reaction mechanisms · CPC title