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High-flux nucleic acid analysis method and application thereof

A nucleic acid analysis, high-throughput technology, applied in the field of biotechnology and molecular diagnosis, can solve the problems of lack of detection methods, quantitative analysis, such as expression and copy number analysis

Active Publication Date: 2014-07-02
GENESKY TECH (SUZHOU) INC +1
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These enrichment techniques are mainly used for mutation detection of candidate genes, but because these enrichment processes eliminate the proportional relationship between the product and the amount of the original template to a certain extent, the quantitative analysis of the enriched candidate gene fragments cannot be accurately realized, such as Expression and copy number analysis
[0011] Therefore, there is still a lack of effective detection methods for gene detection in this field, especially gene identification, gene expression analysis, DNA methylation analysis, mutation screening, SNP typing, CNP typing and CNV detection, so there is an urgent need Development of an efficient method for high-throughput genetic analysis

Method used

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  • High-flux nucleic acid analysis method and application thereof
  • High-flux nucleic acid analysis method and application thereof
  • High-flux nucleic acid analysis method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0150] Detect the typing of 48 SNP sites

[0151] Design junction probes for 48 SNP sites, design 3 probes for each site, 2 5'-end allele-specific probes and 1 3'-end consensus sequence, the first half of the 5'-end probe A general PCR sequence compatible with the next-generation sequencing platform of Illumina is added in part, and another general PCR sequence compatible with the next-generation sequencing platform of Illumina is added to the second half of the 5' end probe. The probe was ligated under the action of TaqDNA ligase under the condition of good pairing with the template, and the ligation product was amplified using general PCR primers compatible with the illumina next-generation sequencing platform. Different samples were amplified with general primers with different tag sequences, and then After uniform mixing and purification, 1x72 sequencing was performed on the Illumina GAIIx sequencer. After the Sequencing reads are read out by software, different sample so...

Embodiment 2

[0180] Detection of DMD gene exon deletion duplication

[0181] Basic principles such as Figure 4 As shown, 141 probes were designed for each sample, 129 of which were distributed on the 79 exons of the DMD gene, 6 reference gene probes, and 6 sex chromosome sex identification probes (3 on the X chromosome, 3 on the Y chromosome). Two probes are designed for each site, one 5' end probe and one 3' end probe. For the specific sequence that hybridizes with the target nucleic acid fragment, the 5' end of the 3' end probe is modified by phosphorylation, the first half is the specific sequence that hybridizes with the target nucleic acid fragment, and the second half is the general sequence consistent with the subsequent PCR amplification primers . The probes were ligated under the action of Taq DNA ligase under the condition of good pairing with the template, and the ligation products were amplified with universal PCR primers compatible with the Illumina next-generation sequenc...

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Abstract

Disclosed are a method for analyzing high-throughput nucleic acid and an application thereof. Among n nucleic acid fragments to be analyzed, for each target nucleic acid fragment, at least two specific probes bound to different binding domains of the target nucleic acid fragment are provided, each specific probe has a specific binding domain and a universal sequence domain, a sequence of the specific binding domain and a sequence of the binding domain of the target nucleic acid fragment are complementary, and a sequence of the universal sequence domain is corresponding to a sequence of a sequencing primer.

Description

technical field [0001] The invention belongs to the field of biotechnology and molecular diagnosis, in particular, the invention relates to a high-throughput nucleic acid analysis method and its application. Background technique [0002] Gene is the material basis of heredity, and it is a specific nucleotide sequence with genetic information on DNA or RNA molecules. Except for some viruses whose genetic material is RNA, the genetic material of almost all non-viral organisms is DNA. Different species have their specific gene sequences, so by detecting the gene sequences in a sample, the biological species in the sample can be judged. [0003] In the process of life, genes are transcribed into mRNA through DNA, and then use mRNA as a template to translate biologically active protein molecules, thereby expressing the genetic information stored in the DNA sequence. By analyzing the amount of each mRNA in different tissues and combining the differences in physiological function...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/68C12Q1/6827C12Q2525/155C12Q2525/161C12Q2535/131C12Q1/6869C12Q2535/122C12Q2521/501C12Q2533/107
Inventor 姜正文杨锋
Owner GENESKY TECH (SUZHOU) INC
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