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Method for screening nucleic acid aptamer

A nucleic acid aptamer, a technology for nucleic acids, used in biochemical equipment and methods, microbial determination/inspection, DNA preparation, etc., can solve problems such as unpredictable changes in electrophoretic mobility and difficulty in applying CE-SELEX, achieving excellent The effect of increasing the acquisition rate and the acquisition rate

Active Publication Date: 2018-11-09
THE UNIV OF TOKYO +1
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Problems solved by technology

In this method of setting the sorting region, there is a problem that a sequence with low binding ability that dissociates halfway is easily included in the sorting region ( figure 1 A)
[0005] In addition, when small molecules such as low-molecular compounds are used as targets, changes in electrophoretic mobility when binding to ssDNA libraries can hardly be expected, so it is considered difficult to apply CE-SELEX

Method used

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  • Method for screening nucleic acid aptamer
  • Method for screening nucleic acid aptamer
  • Method for screening nucleic acid aptamer

Examples

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Embodiment 1

[0049] Immobilization of target molecules on magnetic particles

[0050] Dynabeads MyOne (trademark) Carboxylic Acid (Invitrogen), a magnetic particle having a carboxyl group on the surface, was used as a carrier, and molecules were immobilized according to the operating procedures attached to the product. Add 20mM Tris-HCl, 10mM NaCl, 1mM MgCl 2 100 μl of buffer solution (pH 7.4) was stored at 4° C. as a 10 mg / ml magnetic particle stock solution.

[0051] CE-SELEX condition optimization

[0052] Capillary Electrophoresis Conditions

[0053] A capillary electrophoresis system (Agilent 7100: Otsuka Electronics) was used. As the capillary, a bubble cell fused-silica capillary (Agilent technologies) with a length of 80.6 cm, an effective length (length to the detection window) of 72.2 cm, and an inner diameter of 75 μm was used. The capillary was placed in a cassette so that equal lengths of the capillary were exposed between the electrodes on the input side (injection port, ...

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Abstract

The present invention pertains to a method for screening a nucleic acid aptamer that includes the following steps: (a) bringing a target molecule fixed to a solid-phase carrier into contact with a nucleic acid aptamer candidate; (b) fractionating the nucleic acid aptamer candidate bound to the target molecule via capillary electrophoresis; and (c) amplifying the nucleic acid aptamer candidate viaPCR.

Description

technical field [0001] The present invention relates to a method for screening nucleic acid aptamers. Background technique [0002] The so-called nucleic acid aptamer refers to single-stranded DNA or RNA with molecular recognition ability, which was first reported by Ellington et al. and Tuerk et al. in 1990. Nucleic acid aptamers can be obtained by an evolutionary engineering method called Systematic Evolution of Ligands by Exponential enrichment (SELEX), and a large number of aptamers with binding ability and specificity comparable to antibodies have been reported. Sexual nucleic acid aptamers. In addition, aptamers can be obtained for various targets such as proteins and cells, such as low-molecular compounds that are difficult to obtain antibodies, and their application to therapeutic drugs and diagnostic drugs is expected. However, the acquisition probability of antibodies is said to be 90% or more, whereas the acquisition probability of conventional nucleic acid apta...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/115
CPCC12N15/115C12Q1/6811C12Q2525/205C12Q2531/113C12Q2565/518C12N15/1048C12N2310/16C12Q2541/101
Inventor 吉本敬太郎和久井幸二古性均
Owner THE UNIV OF TOKYO
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