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Method for treating single-stranded DNA, and applications thereof

A DNA pairing and single-strand technology, which is applied in biochemical equipment and methods, microbial measurement/inspection, library creation, etc., can solve the problems of missing single-strand DNA and low utilization rate of sample molecules, and improve the utilization rate of molecules , low detection limit, and the effect of expanding the detection range

Active Publication Date: 2020-02-14
BGI GUANGZHOU MEDICAL LAB CO LTD +2
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

For this reason, an object of the present invention is to propose a method for constructing a library based on single-stranded DNA, which can solve the defect that many single-stranded DNAs are missed during the construction of traditional double-stranded DNA libraries, resulting in low utilization of sample molecules

Method used

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  • Method for treating single-stranded DNA, and applications thereof
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  • Method for treating single-stranded DNA, and applications thereof

Examples

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Embodiment 1

[0044] Embodiment 1 provides a method for constructing a sequencing library based on single-stranded DNA (technical route such as figure 1 shown), the experimental samples used the positive / negative plasma standard HD777 and HD779 purchased from Horizon (standard product numbers match the data shown later). The adapters and sequence primers used in the experiment are shown in Table 1 below.

[0045] Table 1 Primer sequences used in DNA library construction

[0046]

[0047] Wherein, N in Table 1 represents a random sequence, which can be any one of A, T, C, and G. * stands for thio modification, which means thio modification of the base in front of *. Taking the sequence of SEQ ID NO:3 as an example, the first * from left to right (that is, the number from the 5' end to the 3' end) represents, Thio-modify the preceding base. The purpose of thio-modifying the base is to prevent the exolytic activity of the polymerase from digesting the primer. dd stands for dideoxy modif...

Embodiment 2

[0086] Example 2 Library targeted capture

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Abstract

The invention discloses a method for treating single-stranded DNA, and applications thereof, and relates to the field of gene sequencing, particularly to a method for constructing a library based on single-stranded DNA. The method comprises: linking a first linker to the 3' tail end of single-stranded DNA, wherein the first linker is linked to binding agent, and comprises two partially complementary strands, the two partially complementary strands are respectively a first strand and a second strand, and the second strand has 5-10 basic groups more than the first strand; and adsorbing the single-stranded DNA linked to the first linker by using an adsorbent so as to remove the second strand, so that the double-stranded DNA containing the single-stranded DNA is obtained. According to the invention, the gene library constructed by the method has advantages of high molecular utilization rate, low initial quantity of samples for library construction and low minimum detection limit.

Description

technical field [0001] The invention relates to the field of gene sequencing, in particular to a method and application for processing single-stranded DNA. Background technique [0002] The occurrence of malignant tumors is the result of accumulation of multiple genes and multiple events. In the process of clonal evolution, a series of gene mutations are often accumulated, which may involve the changes of multiple genes on different chromosomes, such as oncogenes, tumor suppressor genes, apoptosis genes, cell cycle regulatory genes and maintenance cell genomes. Stability genes, etc. High-throughput sequencing technology can be used to study the mutations and changes in expression levels of the above-mentioned genes and their expressed mRNAs, thereby revealing changes in signaling and metabolic pathways in tumorigenesis and evolution. However, the current high-throughput sequencing library mainly uses double-stranded DNA templates as the starting point, and undergoes steps ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C40B50/06C12Q1/6858
CPCC40B50/06C12Q1/6858C12Q2535/122
Inventor 王晶晶刘继龙宋炎刘磊叶明芝谭美华
Owner BGI GUANGZHOU MEDICAL LAB CO LTD
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