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A psp-based elRNA library construction method suitable for high-throughput sequencing

A library construction, high-throughput technology, applied in the field of elRNA library construction suitable for high-throughput sequencing based on PSP, can solve the problem of affecting the success rate of high-throughput sequencing library construction, inability to perform high-throughput sequencing, and exosomes. problems such as low content, to achieve the effect of easy practical implementation, simple and feasible method, and strong purpose

Active Publication Date: 2019-05-24
江苏吉诺思美精准医学科技有限公司
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Problems solved by technology

[0004] However, the content of exosomes is very low, which has a certain impact on the research of RNA in exosomes using high-throughput sequencing technology, and specifically affects the success rate of library construction in high-throughput sequencing.
In layman's terms, if the library construction is not successful, high-throughput sequencing cannot be performed, and related research cannot be carried out.

Method used

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  • A psp-based elRNA library construction method suitable for high-throughput sequencing
  • A psp-based elRNA library construction method suitable for high-throughput sequencing
  • A psp-based elRNA library construction method suitable for high-throughput sequencing

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Embodiment 1

[0053] Example 1 Technical example of the present invention suitable for high-throughput library construction of extremely low-content elRNA

[0054] 1. Use the 2100 nucleic acid detector to perform quality control on the extracted exosome total RNA. The results are as follows: figure 1 ;

[0055] 2. Add 10ng of exosomal total RNA to the PCR tube;

[0056] 3. Follow the method and steps provided by the library building kit to build the library until the ligation product is purified; the specific steps are shown in (Ⅰ)~(IV):

[0057] (I) Ribosomal RNA was removed from exosomal Total RNA by using a ribosomal RNA removal kit;

[0058] (II) Perform high-temperature and high-salt interruption of the RNA obtained in step (I) using the reverse transcription module buffer provided by the kit;

[0059] (Ⅲ) Reverse transcribe the product obtained in step (II) into double-stranded cDNA using the reverse transcription module provided by the kit, and purify it with magnetic beads;

[0...

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Abstract

The invention discloses a method for constructing an elRNA library based on PSP and applicable to high-flux sequencing. The method comprises the following steps: (1) applying elRNA as a sample source; (2) carrying out library construction by using a ribosome removing method until a PCR reaction is to be carried out; and (3) carrying out PCR amplification twice by using a PSP library enrichment technology. With development of high-flux sequencing technology, sample problems are increasingly severe, and the content of inclusions in RNA coated by exosome, especially in exosome large RNA (i.e., elRNA), is very low; thus, the invention provides the library construction method applicable to elRNAs and based on the PSP library enrichment technology. The method provided by the invention is simple in operation flow and can obtain a library needed in high-flux sequencing.

Description

technical field [0001] The invention relates to the technical field of gene high-throughput sequencing, in particular to a PSP-based method for constructing an eRNA library suitable for high-throughput sequencing. Background technique [0002] In second-generation and third-generation sequencing technologies, constructing high-quality sequencing libraries is a key factor for high-throughput sequencing. The quality of the sequencing library directly determines the quality of the data produced by the sequencing, which in turn has a great relationship with the data analysis. With the rapid development of high-throughput sequencing technology, major reagent companies, high-throughput sequencing has been greatly applied in various fields such as zoology, botany, microbiology, agriculture, and medicine. High-throughput sequencing library construction methods are gradually being developed, matured, and applied; the types of samples faced are becoming more and more complex, such as...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10C12Q1/6806C12Q1/6869C40B50/06
CPCC12N15/1096C12Q1/6806C12Q1/6869C40B50/06C12Q2531/113C12Q2525/191C12Q2535/122
Inventor 曹振龙冷小健刘少卿邢红兵
Owner 江苏吉诺思美精准医学科技有限公司
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