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Construction method for trace frozen tissue ATAC-seq sequencing library

A technique for sequencing libraries and constructing methods, which is applied in the field of molecular biology and can solve problems such as inapplicability

Pending Publication Date: 2021-11-05
四川普罗海尔斯生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In summary, the existing ATAC-seq technology is not suitable for frozen microtissues, especially frozen squamous cell carcinoma microtissues

Method used

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  • Construction method for trace frozen tissue ATAC-seq sequencing library
  • Construction method for trace frozen tissue ATAC-seq sequencing library
  • Construction method for trace frozen tissue ATAC-seq sequencing library

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] A library construction method for ATAC-seq sequencing of frozen microscopic esophageal squamous cell carcinoma tissues.

[0036] (1) Configure the reagents required for ATAC-seq library construction

[0037] a.6X HB unstable

[0038]

[0039] b.1X HB unstable

[0040]

[0041]

[0042] c. 50% Iodixanol Solution

[0043] Reagent Volume of 1 sample 6X HB unstable 75 60% Iodixanol Solution 375

[0044] d. 40% Iodixanol Solution

[0045] Reagent Volume of 1 sample 6X HB unstable 112.5 1M Sucrose 108 60% Iodixanol Solution 450 H2O 4.5

[0046] e. 30% Iodixanol Solution

[0047] Reagent Volume of 1 sample 6X HB unstable 112.5 1M Sucrose 108 60% Iodixanol Solution 337.5 H2O 117

[0048] (2) Grinding tissue and extracting cell nuclei

[0049] a. Pipette 2 mL of pre-cooled 1 x HB unstable buffer into a tissue grinder, place on ice, and set aside.

[0050] b....

Embodiment 2

[0100] A library construction method for ATAC-seq sequencing of frozen microintestinal tissue.

[0101] (1) Configure the reagents required for ATAC-seq library construction

[0102] a.6X HB unstable

[0103]

[0104] b.1X HB unstable

[0105]

[0106]

[0107] c. 50% Iodixanol Solution

[0108] Reagent Volume of 1 sample 6X HB unstable 75 60% Iodixanol Solution 375

[0109] d. 40% Iodixanol Solution

[0110] Reagent Volume of 1 sample 6X HB unstable 112.5 1M Sucrose 108 60% Iodixanol Solution 450 H2O 4.5

[0111] e. 30% Iodixanol Solution

[0112] Reagent Volume of 1 sample 6X HB unstable 112.5 1M Sucrose 108 60% Iodixanol Solution 337.5 H2O 117

[0113] (2) Grinding tissue and extracting cell nuclei

[0114] a. Pipette 2 mL of pre-cooled 1 x HB unstable buffer into a tissue grinder, place on ice, and set aside.

[0115] b. Quickly weigh a small amount ...

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Abstract

The invention provides a construction method for a trace frozen tissue ATAC-seq sequencing library. The method comprises the following steps of S1, grinding tissues, preparing a cell suspension and extracting cell nucleuses; S2, carrying out transposition and purification; S3, carrying out PCR enrichment, constructing the library and screening fragments; and S4, carrying out library quality inspection. The construction method for the micro frozen tissue ATAC-seq sequencing library has the following advantages that the method can well solve the problem that an existing ATAC-seq technology is not suitable for frozen micro tissues, especially frozen squamous cell carcinoma micro tissues.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a method for constructing an ATAC-seq sequencing library of microfrozen tissue samples. Background technique [0002] Chromatin transposase accessibility sequencing (assay for transposase-accessible chromatin with high throughput sequencing, ATAC-seq) is a new technology for chromatin transposase accessibility based on high-throughput sequencing. It was first published in Nature Methods in 2013 and developed by William JGreenleaf's research group at Stanford University. The principle is to use the property of the transposase Tn5 to cut the open chromatin region, and sequence the DNA sequence captured by the Tn5 enzyme. Compared with previous chromatin openness research techniques, ATAC-seq requires less samples, the process is simpler, and the efficiency is higher. It only takes two steps to capture open chromatin regions from a small number of cells. This...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806C12Q1/6869C40B50/06
CPCC12Q1/6806C12Q1/6869C40B50/06
Inventor 姜昊邓昭敏
Owner 四川普罗海尔斯生物科技有限公司