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Novel DNA library construction kit and application thereof

A kit and reagent technology, applied in the field of molecular biology, can solve the problems of ineffective utilization of DNA information, loss of DNA information, unsatisfactory sample amplification at breakpoints, etc.

Inactive Publication Date: 2018-06-15
SHANGHAI MAJORBIO BIO PHARM TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the PCR amplification method is biased towards fragments, and generally can only cover areas with better quality, and the amplification situation of samples with breakpoints is very unsatisfactory, and all DNA information in the samples cannot be effectively used; while in double-stranded In the ligation scheme, the efficiency of adapter ligation is generally poor in regions that are greatly affected by library fragmentation, which will also lead to the loss of DNA information

Method used

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  • Novel DNA library construction kit and application thereof
  • Novel DNA library construction kit and application thereof
  • Novel DNA library construction kit and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0213] 1. Preparation of DNA library construction kit

[0214] (1) Synthesis of cohesive end adapters in the DNA library construction kit:

[0215] The sticky end adapter is a double-stranded linker with a sticky end, one end of the sticky end adapter is a first sticky end, and the first sticky end includes at least a random sequence.

[0216] 1. As an example, use Primer1 and Primer2 to synthesize sticky end adapters,

[0217] Primer1: 5'-AGATCGGAAG-3' (SEQ ID NO.1);

[0218] Primer2: 5'-CTTCCGATCTNNNNNN-3' (SEQ ID NO.2); wherein, NNNNNN represents a random sequence, usually the length of the random sequence is 2-30nt, preferably 4-20nt, more preferably 5-10nt .

[0219] 2. Add the configuration reaction solution according to Table 1:

[0220] Table 1

[0221] Reagent

Volume (ul)

Primer1 (10μM)

1

Primer2 (10μM)

1

TE buffer

7

5M NaCl

1

total capacity

10

[0222] The TE buffer used in the reaction can als...

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Abstract

The invention relates to the technical field of molecular biology and in particular relates to a novel DNA library construction kit and an application thereof. The DNA library construction kit provided by the invention comprises an adhesive end joint, a ligase, a DNA extension reagent and a DNA amplification reagent. The DNA library construction kit disclosed by the invention can be applied to a sample with low initial mass and can directly mediate the connection of DNA and the joint by a common ligase; and compared with previous methods, the kit has the advantages that the connection efficiency of samples is greatly increased, the diversity of the samples are retained to the greater extent, the stability is good, and library bias caused by artificial factors is shortened.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a novel DNA library construction kit and its application. Background technique [0002] The function of DNA molecules is to store all the genetic information of almost all proteins and RNA molecules that determine the traits of species. The interpretation of DNA information is of great significance for us to study the mysteries of life and the mechanism of diseases. DNA molecules usually exist in samples in the form of double strands, but the vast majority of precious medical samples exist in the form of formalin-fixed paraffin-embedded (FFPE) samples. [0003] FFPE samples are precious, but there are difficulties in amplification in molecular biology experiments. Usually FFPE is of poor quality, fragmented and has many breakpoints on the DNA, which poses certain difficulties for the amplification of the corresponding samples, because usually the polymerase cannot skip...

Claims

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Application Information

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IPC IPC(8): C40B50/06C12N15/10C12Q1/6869
CPCC12N15/1093C12Q1/6869C40B50/06C12Q2525/191C12Q2521/501C12Q2533/101
Inventor 陈昌岳王芳李静李明明查洋胡秋萍张祥林
Owner SHANGHAI MAJORBIO BIO PHARM TECH
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