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Small-fragment DNA library construction method capable of improving Hi-C library data quality

A DNA library and data quality technology, applied in the field of gene sequencing, can solve the problems of information loss, low effective data rate of capture method, high random noise of Hi-C interactive data, etc., to achieve cost reduction, success rate improvement, and operation process simple effect

Pending Publication Date: 2020-07-28
嘉兴菲沙基因信息有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, it is still far away from the goal of the research to reveal the biological function generally, and the Hi-C interaction data contains quite high random noise
At the same time, the experiment found that the construction of the existing Hi-C high-throughput sequencing library also has the problems of information loss and a certain preference in the capture method, resulting in a low effective data rate.

Method used

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  • Small-fragment DNA library construction method capable of improving Hi-C library data quality
  • Small-fragment DNA library construction method capable of improving Hi-C library data quality
  • Small-fragment DNA library construction method capable of improving Hi-C library data quality

Examples

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Embodiment 1

[0036] In this example, the Hi-C ligation product is used as the starting material, and it is necessary to provide a method for preparing the Hi-C ligation product of mouse embryonic stem cells in the prior art.

[0037] This embodiment provides a method for constructing a small fragment DNA library of mouse embryonic stem cells Hi-C, the method specifically comprising the following steps:

[0038] 1. Target fragment recovery

[0039] 1) Refer to the operation instructions of DNeasy Blood&Tissue Kit and modify some steps;

[0040] 2) Centrifuge the Hi-C ligation product, discard the supernatant, add 180 μL ATL and 20 μL Proteinase K to the sample tube, de-crosslink at 56°C for 2 h, invert the centrifuge tube 2-3 times during the process, and mix the samples;

[0041] 3) Add 200 μL of buffer AL to mix well, incubate at 56°C for 10 min, invert the centrifuge tube 2-3 times during the process, and mix the samples;

[0042] 4) Add 200 μL of 96-100% ethanol and mix well;

[0043] ...

Embodiment 2

[0109] In this example, the DNA of the Hi-C ligation product is used as the starting material, and it is necessary to provide a method for preparing the DNA of the Hi-C ligation product of zebrafish liver cells in the prior art.

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Abstract

The invention discloses a small-fragment DNA library construction method capable of improving Hi-C library data quality. The method specifically comprises the following steps: S1, firstly, recoveringHi-C ligation product cell nucleuses by using DNeasy Blood & Tissue Kit or purifying existing Hi-C ligation product DNA; S2, performing fragmentation, terminal repairing and A addition on recovered fragments to obtain terminal-repaired DNA; S3, performing Index connection on the terminal-repaired DNA to obtain a connected material; S4, purifying the connected material to obtain a DNA purified material; and S5, finally, performing PCR amplification on the DNA purified material to obtain a DNA sequencing library. The invention relates to the technical field of gene sequencing. According to the small-fragment DNA library construction method capable of improving Hi-C library data quality, existing DNA small-fragment library construction experiment technology is subjected to a plurality of optimizations, so a library construction success rate is substantially improved, and cost is significantly reduced; and the operation process of the method is simple, and the method can be copied to otherlaboratories having molecular biology bases.

Description

technical field [0001] The invention relates to the technical field of gene sequencing, in particular to a small fragment DNA library construction method which can improve the data quality of Hi-C library. Background technique [0002] High-throughput sequencing technology, also known as next-generation sequencing technology, is marked by the ability to sequence hundreds of thousands to millions of DNA molecules in parallel and with shorter read lengths. The second-generation high-throughput illumina sequencing platform has It has the advantages of high throughput, high accuracy and low cost, and is widely used in many fields. [0003] Chromosome Conformation Capture (3C) technology is a technique for studying chromosome and protein interactions and chromosome conformation, which can provide detailed information on the association between distant genetic loci, which can be obtained from formaldehyde-fixed captured in the nucleus and can be inferred from the three-dimensiona...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C40B50/06C40B40/06
CPCC40B50/06C40B40/06
Inventor 张骥诚
Owner 嘉兴菲沙基因信息有限公司
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