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Method for constructing single-stranded annular library

A single-stranded circular library and construction method technology, applied in chemical libraries, biochemical equipment and methods, microbial measurement/testing, etc., can solve the problems of reduced sequencing quality and efficiency, affecting the combination of sequencing primers, and cumbersome operations , to achieve the effect of shortening the database construction cycle, improving the efficiency of database construction, and good compatibility

Active Publication Date: 2019-05-14
MGI TECH CO LTD
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Problems solved by technology

When the circular single-stranded library is sequenced downstream, the ME will be annealed and complementary, which will affect the binding of the sequencing primers and significantly reduce the quality and efficiency of the sequencing.
In addition, the experimental operation of this method also requires operations such as gap tr

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  • Method for constructing single-stranded annular library
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  • Method for constructing single-stranded annular library

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Embodiment 1

[0085] Example 1, construction of single-stranded circular library

[0086] In this example, 50ng of the genome was used for testing, and the construction method of the single-stranded circular library is as follows:

[0087] 1. Synthesize a biotin-containing adapter sequence (i.e., single-stranded DNA). The adapter sequence includes three types of a1, b1, and c1, wherein c1 is the common sequence of a1 and b1 (i.e., the recognition sequence of TN5 transposase, ME sequence) complementary sequence.

[0088] a1: (Sequence 1 in the sequence table, the 1-16th (i.e. bold part) of sequence 1 is recorded as sequence A, the underlined region is the recognition sequence of transposase TN5, and biotin is used as a capture tag for the following single-stranded DNA capture);

[0089] b1: AGATGTGTATAAGAGACAG;

[0090] c1: CTGTCTCTTATACACATCT.

[0091] 2. After diluting a1, b1 and c1 in step 1 to 200 μM, mix a1 and c1 in equal amounts to obtain an a1-c1 mixture; mix b1 and c1 in equal...

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Abstract

The invention discloses a method for constructing a single-stranded annular library. The method for constructing the single-stranded annular library comprises the following steps: 1) a target DNA is interrupted by utilizing a transposase complex to obtain a DNA disruption product; the transposase complex comprises transposase, a linker A and a linker B; the linker A consists of a single-stranded DNA and a complementary sequence of a recognition sequence, wherein the single-stranded DNA is obtained by sequentially connecting a capture label, the 1st to 16th positions of a sequence 1 in a sequence table and the recognition sequence of the transposase; the linker B consists of the recognition sequence and the complementary sequence of the recognition sequence; 2) notch translation and meltingare carried out on the DNA disruption product, and capturing is carried out by using the capture label to obtain the single-stranded DNA containing the capture label; 3) the single-stranded DNA containing the capture label is cyclized to obtain the single-stranded annular library. The method for constructing the single-stranded annular library has the advantages of simple process and simple operation, does not need to carry out operations such as end repair, linker addition, single-stranded separation and the like, can shorten the library building period, and can improve the library buildingefficiency.

Description

technical field [0001] The invention relates to a method for constructing a single-chain circular library in the field of biotechnology. Background technique [0002] Whole genome sequencing is the sequencing of the genomes of individuals of species of unknown genome sequence. The more common whole-genome construction library is a double-stranded DNA library based on the Illumina platform or Proton platform. The test process is roughly as follows: Genomic DNA is randomly broken into 180-280bp fragments by physical or chemical methods, and the ends are repaired and A is added. After the tail, adapters are connected to both ends of the fragments, and after linear amplification by chain polymerase reaction (PCR), quality inspection is performed, and sequencing can be carried out after passing the test. [0003] The single-stranded circular library is a closed single-stranded circular library with a linker sequence, and its main application platforms include the Complete Genomi...

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Application Information

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IPC IPC(8): C12Q1/6806C12N15/10C40B50/06
Inventor 严海生张通达郭健
Owner MGI TECH CO LTD
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