Multi-nucleic acid co-labeling support, preparation method therefor, and application thereof

A nucleic acid labeling and support technology, applied in biochemical equipment and methods, nucleotide libraries, library creation, etc., can solve problems such as inability to change liquids, unstable suspension characteristics of droplets, and cell RNA diffusion out of holes

Pending Publication Date: 2022-02-25
BEIJING SEEKGENE BIOSCIENCES CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The droplet method completely isolates cells and label microbeads from other cells and microbeads through water-in-oil, effectively reducing the possibility of cross-contamination; at the same time, in addition to realizing the 3' RNA expression profile library, the droplet scheme can also separate cells Tags and molecular tags are coupled with template-switching sequences to achieve 5' single-cell RNA expression profile sequencing; however, due to the instability and suspension characteristics of the droplets themselves, the single-cell library construction scheme based on the droplet method is difficult to obtain when the RNA is captured by cells. The medium ca

Method used

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  • Multi-nucleic acid co-labeling support, preparation method therefor, and application thereof
  • Multi-nucleic acid co-labeling support, preparation method therefor, and application thereof
  • Multi-nucleic acid co-labeling support, preparation method therefor, and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0097] Example 1: A variety of nucleic acid co-labeled supports are applied to the construction of 5' single-cell RNA expression profile library and VDJ library construction and multi-omics library construction

[0098] In this example, various nucleic acid co-labeled supports were prepared according to the following steps and applied to construct 5' single-cell RNA expression profile library, VDJ library and multi-omics library.

[0099] 1Making a variety of nucleic acid-labeled magnetic beads

[0100] 1.1 Synthesize a single-stranded nucleic acid of the following sequence.

[0101]

[0102] 1.2 At a concentration of 0.25M EDC, 384 kinds of 300pmol amino-modified CB1 single-stranded nucleic acid (SEQ ID No.2) and 300pmol amino-modified dT single-stranded nucleic acid (SEQ ID No.1) and 60,000 30μM carboxyl magnetic beads were placed at room temperature Rotate and mix for 3 hours, and obtain 384 kinds of nucleic acid-labeled magnetic beads after washing twice.

[0103] 1.3...

Embodiment 2

[0136] Example 2: A variety of nucleic acid co-labeled supports are applied to the construction of 3' single-cell RNA library

[0137] In this example, multiple nucleic acid co-labeled supports were prepared according to the following steps and applied to construct a 3' single-cell RNA library.

[0138] 1Making a variety of nucleic acid-labeled magnetic beads

[0139] 1.1 Synthesize a single-stranded nucleic acid of the following sequence.

[0140]

[0141] 1.2 At 0.25M EDC concentration, 384 kinds of 300pmol amino-modified CB1 single-stranded nucleic acid (SEQ ID No.2) and 300pmol amino-modified SP2-dT30VN single-stranded nucleic acid (SEQ ID No.11) and 60,000 30μM carboxyl magnetic The beads were rotated and mixed at room temperature for 3 hours, and 384 kinds of nucleic acid-labeled magnetic beads were obtained after washing twice.

[0142] 1.3 Mix the 384 kinds of nucleic acid-labeled magnetic beads obtained in 1.2 and distribute them evenly into 384-well plates.

[...

Embodiment 3

[0170] Example 3: A variety of nucleic acid co-labeled supports are applied to the construction of single-cell transcriptome library

[0171] In this example, multiple nucleic acid co-labeled supports were prepared according to the following steps and applied to construct a single-cell transcriptome library.

[0172] 1Making a variety of nucleic acid-labeled magnetic beads

[0173] 1.1 Synthesize a single-stranded nucleic acid of the following sequence.

[0174] SEQ ID No. name sequence 14 CB2-dN6 CTACAGnnnnnnnnNNNNNN

[0175] 1.2 At a concentration of 0.25M EDC, 384 kinds of 300pmol amino-modified CB1 single-stranded nucleic acid (SEQ ID No.2) and 300pmol amino-modified dT single-stranded nucleic acid (SEQ ID No.1) and 60,000 30μM carboxyl magnetic beads were placed at room temperature Rotate and mix for 3 hours, and obtain 384 kinds of nucleic acid-labeled magnetic beads after washing twice.

[0176] 1.3 Mix the 384 kinds of nucleic acid-labeled ...

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Abstract

Provided are a variety of nucleic acid co-labeling supports, a preparation method therefor, and an application thereof. The supports comprise a support body, and a variety of nucleic acid labels located on the surface and/or inside of the support body. The nucleic acid labeled on a single support at least comprises: one or more first nucleic acid labels having a function that at least comprises of trapping a specific compound in a reaction system onto the surface of the support; and one or more second nucleic acid labels having a function that at least comprises participating in a specified biochemical reaction process of the specific compound trapped onto the surface of the support. The foregoing variety of nucleic acid co-labeled supports can be used for 5'-terminus single-cell RNA expression profile analysis, construction of a 5'-terminus single-cell VDJ library for a microwell array platform, construction of a 3'-terminus single-cell RNA library, construction of a single-cell transcriptome library, single-cell multi-omics research, multiplex PCR and/or construction of a multiplex PCR sequencing library, etc.

Description

technical field [0001] The invention relates to a multi-nucleic acid co-labeling support and its production method and application. Background technique [0002] Traditional nucleic acid molecular reactions, including nucleic acid hybridization, extension, amplification and other reactions, are all carried out in the liquid phase. The liquid phase provides a uniform and stable environment for the nucleic acid and enzyme reactions involved in the reaction to maximize the output. With the deepening of biological research, scientists have discovered that linking the nucleic acid or enzyme involved in the reaction to the solid-phase surface can give the nucleic acid spatial position information to facilitate purification, separation, detection and analysis. Therefore, more and more solid-phase nucleic acids have been developed. The reaction is used for nucleic acid sequence analysis and nucleic acid quantification, such as nucleic acid chip technology where oligonucleotides are ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6806C40B40/06C40B50/06
CPCC12Q1/6806C40B40/06C40B50/06C12Q2563/149C12Q1/68C12N15/11
Inventor 焦少灼韩金桓李研刘书杰马兴勇罗云超桑国芹谢莹莹徐猛李宗文
Owner BEIJING SEEKGENE BIOSCIENCES CO LTD
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