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Method for eliminating self-connector of sequencing library and application

A technology for sequencing libraries and next-generation sequencing libraries, which is applied in the field of self-connector elimination of sequencing libraries, can solve problems such as high pollution of adapter dimers, affecting library output, failing to meet sequencing requirements, etc., achieving obvious changes in adapter content, Improve data efficiency and increase the effect of cleanreads

Pending Publication Date: 2021-12-21
上海金匙医学检验实验室有限公司
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Problems solved by technology

However, reducing the adapter content and adjusting the amount of magnetic beads will affect the output of the library, which cannot meet the requirements of the machine. For samples with a very small sample size, if the sequencing shows high contamination of the adapter dimer, it means that the experiment fails and causes sample loss.
2. Library level, use the library from the library: the library is reamplified and recovered. In the case of extremely high adapter content (>80%), the library amplification efficiency is low, and the proportion of target fragments is still low, which cannot meet the sequencing requirements. Therefore, there is an urgent need for a method that can effectively eliminate adapter dimers at the library level, which can meet the detection requirements while reducing the adapter content.

Method used

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  • Method for eliminating self-connector of sequencing library and application
  • Method for eliminating self-connector of sequencing library and application

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0096] Embodiment 1 The design optimization of this application

[0097]As mentioned in the background technology of the present application, the existing sequencing library, especially the next-generation sequencing library, has the problem of joint interference in the construction process. Under normal circumstances, it is more to consider re-extracting and building a library. Even if re-extracting and constructing a library cannot solve this problem, everyone will think that there is a problem with the quality of the sample; or for the problem of adapters, the conventional method in the next-generation sequencing technology process is still the adapter content. The adjustment and purification of magnetic beads, or the recovery of gel cutting, will not consider processing from the dimension of the library. After all, such samples are rare, especially in scientific research samples, and may be common in clinical samples, but it will not cost A lot of effort has been devoted t...

Embodiment 2

[0124] Example 2 compares the effect with unprocessed library data

[0125] 1. Library screening

[0126] The characteristics of clinical samples are complex, and the extraction and separation efficiency of different samples and the degree of genome fragmentation and degradation affect the recovery rate of the sequence. Some samples with poor quality have a high content of library adapters, and the effective data rate of sequencing is low, which cannot reach the data analysis. requirements.

[0127] From the off-machine data of the illumina sequencing platform, select a library with a normal nucleic acid concentration and a normal concentration of the library, which meets the standards for the Illumina platform, but the output data efficiency is low, and the adapter ratio is as high as 95%. A library with a high concentration of nucleic acid Low, the concentration of the library is normal, which meets the standard of the Illumina platform, but the adapter ratio is 88.5% of th...

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Abstract

The invention relates to a method for eliminating a self-connector of a sequencing library and an application. The method comprises the following steps that a short sequence guide DNA is designed by taking a sequencing connector self-connecting library sequence as a target sequence, double-strand breakage of a dsDNA library molecule is realized by combining with Argonaute endonuclease, and the self-connector is cut from the dsDNA library molecule, so as to prevent the self-connector from being amplified in a subsequent PCR reaction. According to the method for eliminating the self-connector of the sequencing library, the proportion of self-connectors of the library can be remarkably reduced, the sequencing clean reads can be increased, and the data efficiency can be improved.

Description

technical field [0001] The present application relates to the technical field of gene sequencing, and specifically relates to a method and application for eliminating the self-connector of a sequencing library. [0002] technical background [0003] In high-throughput sequencing technology, the quality of the library is crucial to the quality of the data produced by high-throughput sequencing. Low-quality libraries lead to too many Clusters or multiple templates, and the data quality is not high; The coverage rate is low, so the quality of the library directly affects the sequencing effect. The quality of the sequencing library is the key to obtaining high-quality nucleic acid sequencing data. [0004] Low-quality libraries generally refer to libraries with low concentration, dimer pollution, small fragment pollution, large fragment pollution, and too wide peak map, which will lead to low effective data output of the entire lane library and affect the ratio of clean reads. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6806C12Q1/6869C40B50/06
CPCC12Q1/6806C12Q1/6869C40B50/06C12Q2525/191C12Q2521/301
Inventor 曹德盼李东东赵翊婷程珂燕余盼贾雪峰蒋智
Owner 上海金匙医学检验实验室有限公司
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