Buffer solution and application thereof

A buffer solution and divalent cation technology, applied in the field of buffer solution and nucleic acid library construction, can solve the problems of low transformation efficiency, poor compatibility, and difficulty in compatibility with library construction steps.

Active Publication Date: 2021-06-25
ZHEJIANG ANNOROAD BIO TECH CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The existing buffer system has poor sample compatibility and is difficult to be compatible. Different library construction steps require frequent buffer replacement, prolonging the time for library construction, and low conversion efficiency.
[0004] Therefore, the buffer system in the existing library construction needs to be improved

Method used

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  • Buffer solution and application thereof
  • Buffer solution and application thereof
  • Buffer solution and application thereof

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0048] In this example, the buffer of the embodiment of the present invention and the commercially available blue buffer are respectively used as a universal buffer to construct a DNA library, and the method is as follows:

[0049] 1. Procedures and conditions for constructing DNA library:

[0050] (1) Break end repair and adenylate tailing

[0051]

[0052] Wherein, the universal buffer is respectively the aforementioned buffer of the present invention and the commercially available blue buffer, wherein, the buffer formula of the embodiment of the present invention is 5-25mM Tris buffer, 5-20mM divalent cation, 75-150mM- cations, 0.05-2% surfactant and 0.5-2mM dithiothreitol (DTT), pH8-10.

[0053] (2) Add joints

[0054]

[0055] Wherein, the connection buffer is a quick connection buffer (provided by Novizyme Biotechnology Company).

[0056] (3) Purification: Purify with 1× magnetic beads, and perform PCR after elution with an appropriate amount of eluent.

[0057...

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Abstract

The invention discloses a buffer solution and application thereof. The buffer solution comprises 5-25 mM of a Tris buffer solution, 5-20 mM of divalent cations, 75-150 mM of monovalent cations, 0.05-2% by mass of a surfactant and 0.5-2 mM of dithiothreitol, and has a pH value of 8-10. The buffer solution is good in compatibility, high in stability and suitable for multi-step reactions in a library construction process, enables multi-step reactions in the library construction process to be continuously carried out, does not purification, remarkably shortens reaction time, improves the efficiency and stability of a method for constructing a DNA library, and has no obvious inhibition effect on T4DNA ligase.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to a buffer and its application, more specifically to a buffer, a kit, the buffer used in the construction of a nucleic acid library as a general buffer, and a construction Methods for Nucleic Acid Libraries. Background technique [0002] Next-generation sequencing of genomic DNA and corresponding bioinformatics analysis have been widely used in the fields of medical health and technology services. [0003] The general process for the construction of a next-generation sequencing library is as follows: fragment the target DNA; blunt the fragmented DNA; protrude adenylation at the 3' end of the blunt DNA; The DNA fragments are ligated with double-stranded Y adapters that highlight thymine. The existing buffer system has poor sample compatibility and is difficult to be compatible with different library construction steps. Frequent buffer replacement is required, which prolongs th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C40B50/06
CPCC40B50/06
Inventor 潘伟业程世月王亚蕾李志民李大为玄兆伶王海良王娟
Owner ZHEJIANG ANNOROAD BIO TECH CO LTD
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