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Single-stranded DNA, G-quadruplex and preparation method and application of G-quadruplex

A tetramer, single-chain technology, which is applied in the preparation of sugar derivatives, chemical instruments and methods, and medical preparations of inactive ingredients, etc., to achieve the effects of high-efficiency synthesis, easy promotion and application, and simple operation.

Active Publication Date: 2020-09-22
HUNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, there are few reports on the hydrophobic modification of G-tetramer nanostructures to construct biomimetic nanochannels and regulate molecular ion transport on phospholipid membranes.

Method used

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  • Single-stranded DNA, G-quadruplex and preparation method and application of G-quadruplex
  • Single-stranded DNA, G-quadruplex and preparation method and application of G-quadruplex
  • Single-stranded DNA, G-quadruplex and preparation method and application of G-quadruplex

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Embodiment 1

[0034] A kind of G-tetramer of the present invention uses the single-stranded DNA rich in guanine G base as the carrier, and the 5' end and the middle of the single-stranded DNA are modified with 3 long chains of hydrophobic carbon twelve, at 3 A cholesterol molecule is modified at the 'end, and the sequence of the single-stranded DNA rich in guanine G bases is shown in SEQ ID NO.1, specifically:

[0035] PS2.M: 5'-GTGGGTAGGGCGGGTAGGGTTGG-3'.

[0036] The modified single-stranded DNA is:

[0037] lipoPS2.M: 5'- / SpacerC12 / GTGGGT / SpacerC12 / AGGGCGGGT / SpacerC12 / AGGGTTGGChol-3'.

[0038] Spacer C12 in the sequence is a carbon twelve long chain; Chol is cholesterol.

[0039] The single-stranded DNA forms a highly stable two-dimensional G-tetramer with hydrophobic properties under the action of potassium ions. The center of the G-tetramer structure is a hollow cavity that can serve as a potassium ion-specific recognition channel for controllable ion transport. Incubating the amph...

Embodiment 2

[0046] A G-tetramer of the present invention uses single-stranded DNA rich in guanine G bases as a carrier, and two hydrophobic carbon twelve long chains are modified in the middle of the single-stranded DNA, and one is modified at the 3' end Cholesterol molecules, FAM fluorescent dyes modify the 5' end of DNA single strands. The sequence of the single-stranded DNA rich in guanine G bases is shown in SEQ ID NO.1, specifically:

[0047] PS2.M: 5'-GTGGGTAGGGCGGGTAGGGTTGG-3'.

[0048] The modified single-stranded DNA is:

[0049] lipoPS2.M: 5'-FAMGTGGGT / SpacerC12 / AGGGCGGGT / SpacerC12 / AGGGTTGG Chol-3'.

[0050] Spacer C12 in the sequence is a carbon twelve long chain; Chol is cholesterol.

[0051] A preparation method of the biomimetic potassium ion nanochannel of the present embodiment 1, comprising the following steps:

[0052] (1) Single-stranded DNA is synthesized by solid-phase phosphite triester method. During the synthesis process, FAM dye, cholesterol (Chol), and carbon...

Embodiment 3

[0057] A kind of application of the G-tetramer of embodiment 1 of the present invention in transporting potassium ions as a biomimetic potassium ion nanochannel, its application method comprises the following steps:

[0058] (1) Synthesis of LUV vesicles using the lipid membrane hydration method:

[0059] 1.1. Mix the phospholipid solution containing 30 μM DPPC and 5.1 μM cholesterol evenly, then spin dry on a rotary evaporator at 60°C and 200 rpm to make a thin layer of phospholipid film adhere to the wall of the round bottom flask.

[0060] 1.2. Place the round-bottomed flask containing the phospholipid membrane under vacuum at 0°C for 5 hours, then add 1 mL of 10mM Tris-HClbuffer (containing 0.5mM HPTS and 100mM K + ) Slowly inject the liquid along a certain point of the mouth of the round bottom flask so that the liquid does not pass through the phospholipid film on the bottle wall, suspend and stir for 1 hour, ultrasonicate in a water bath for 15 minutes, and then freeze ...

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Abstract

The invention discloses a single-stranded DNA, a G-quadruplex and a preparation method and application of the G-quadruplex. The G-quadruplex takes single-stranded DNA as a carrier, cholesterol, C12 long chain and / or phospholipid molecules are coupled to the single-stranded DNA, and the G-quadruplex with a secondary structure is formed under the action of potassium ions. The preparation method comprises the following steps: coupling cholesterol, a C12 long chain and / or phospholipid molecule to a base of a single-stranded DNA in a chemical cross-linking manner to obtain a modified single-stranded DNA; and reacting the modified single-stranded DNA with potassium ions to form a lipophilic secondary structure G-quadruplex nanostructure. The lipophilic group and the hydrophobic group are coupledto the base of the single-stranded DNA, so that the high membrane permeability and hydrophobicity of the channel structure are remarkably enhanced, the G-quadruplex nanostructure can be embedded in aphospholipid bilayer and stably exist, and specific ions or molecules are carried along the concentration gradient from bottom to top or from top to bottom; and the G-quadruplex nanostructure can beapplied to construction of a bionic nano-channel for transmembrane transportation of potassium ions.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a preparation method and application of single-stranded DNA, G-tetramer and G-tetramer. Background technique [0002] The G-quadruplex structure is a highly folded and stable secondary structure formed by single-stranded DNA rich in guanine G bases under the action of certain ions or molecules. The G-tetramer structure has a nanometer-scale hollow chamber, which allows some specific ions with a particle size of micronanoscale to combine with water molecules, so as to pass through the channel from bottom to top or top to bottom. [0003] However, there are few reports on the hydrophobic modification of G-tetramer nanostructures to construct biomimetic nanochannels and regulate molecular ion transport on phospholipid membranes. Modification of lipophilic cholesterol, carbon twelve and other hydrophobic groups on the single-stranded DNA rich in G bases can enhance the high hydrophobici...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07H21/04C07H1/00A61K47/28A61K47/26A61K9/127
CPCC07H21/04C07H1/00A61K9/1271A61K47/26A61K47/28
Inventor 刘剑波李春影羊小海王柯敏
Owner HUNAN UNIV
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