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Large-fragment DNA cyclization connection method

A circularization and fragmentation technology, applied in the field of genetic engineering, can solve problems such as unusable, high cost, and complexity

Active Publication Date: 2022-07-22
XUKANG MEDICAL SCI & TECH (SUZHOU) CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although these studies can also effectively improve the connection efficiency, complex and high-standard equipment may be required to produce modified enzymes, or they cannot be used due to patent protection, or commercial reagents need to be purchased at a relatively high price

Method used

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Embodiment 1

[0234] Preparation of cyclized linkers

[0235] A circularized linker is provided, which includes a sense strand (SEQ ID NO: 5) and an antisense strand (SEQ ID NO: 2), both of which are phosphorylated at the 5' end of the sense strand and the 5' end of the sense strand The second base is a U base, and the linker sequence is shown in Table 1 below:

[0236] Table 1

[0237] name Primer sequence (5'-3') SEQ ID NO: 5 5'P-AUAGATGTGTA(Biotin)TAAGAGACAG-3' SEQ ID NO: 2 5'P-CTGTCTCTTATACACATCT-3'

[0238] Experiments were performed using the prepared cyclized linkers.

[0239] Step 1: Genomic DNA Extraction

[0240] Take 1ml of whole blood from healthy people, and extract genomic DNA using a genomic DNA extraction kit (manufacturer: Kangwei Century Item No.: CW2087). No obvious degradation of DNA was detected by 0.8% agarose gel electrophoresis.

[0241] Step 2: Joint Annealing

[0242] Prepare the reaction solution as follows:

[0243] ...

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Abstract

The invention relates to a large-fragment DNA cyclization connection method, application of the large-fragment DNA cyclization connection method in construction of a sequencing library, a cyclization joint and a kit for realizing the method and the application, and a DNA molecule and the sequencing library formed by the cyclization joint and the kit.

Description

technical field [0001] The present invention belongs to the field of genetic engineering, and more particularly relates to a large fragment DNA circular ligation method and its application in the construction of a sequencing library, as well as a circular linker and a kit for realizing the method and application, and the formation thereof. DNA molecules and sequencing libraries. Background technique [0002] In next-generation sequencing, large fragment library construction is an important and basic experimental technique. For species whose genome sequence is unknown or has no genomic information of closely related species, gene mapping by next-generation sequencing is one of the ways to quickly understand a species. In addition, next-generation sequencing also has important application value in the current screening and detection of chromosomal structural variation in the human genome. [0003] Mate-Pair sequencing (MP sequencing) is one of the most commonly used next-gen...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12P19/34C40B50/06
CPCC12N15/111C12P19/34C40B50/06
Inventor 任军高芳芳张鹏
Owner XUKANG MEDICAL SCI & TECH (SUZHOU) CO LTD
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