Reagent kit and method for detecting human alpha defensin 1/3 gene copy number

A technology of gene copy number and defensin, applied in the field of biological diagnosis, can solve the problems of high cost and long time

Inactive Publication Date: 2012-07-11
ZHEJIANG UNIV
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  • Abstract
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Problems solved by technology

[0005] The invention provides a kit for detecting the copy number of human α-defensin 1 / 3 gene, which solves the problems of long time-consuming and high cost of traditional methods

Method used

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  • Reagent kit and method for detecting human alpha defensin 1/3 gene copy number
  • Reagent kit and method for detecting human alpha defensin 1/3 gene copy number
  • Reagent kit and method for detecting human alpha defensin 1/3 gene copy number

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Embodiment Construction

[0030] A kit for detecting the copy number of DEFA1 / DEFA3, the main reagents include: specific fluorescent quantitative PCR primers (4μM / μl) for detecting DEFA1 / DEFA3 sites; specific DEFA1 / DEFA3 probes; internal reference gene ALB specific fluorescent quantitative PCR primer (4μM / μl); specific ALB probe; check sample DNA; positive control DNA; fluorescent quantitative PCR reaction solution.

[0031]According to the corresponding sequences of the internal reference gene ALB and the target gene DEFA1 / DEFA3 in GenBank (number: ALB is NM000477.5, DEFA1 is NM004084.3, DEFA3 is NM005217.3), select a conserved fragment with a length of 50-150bp, and use Primer Express 2.0 The software designs specific probes and primers. The specific probe length is between 16-25 bases, the GC content is 30-80%, the C content is higher than the G content, and the annealing temperature (Tm value) is 65-70°C, avoiding stable dimers and hairpins structure. Design 3-4 pairs of suitable intron-specific ...

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Abstract

The invention discloses a reagent kit and a method for detecting human alpha defensin 1/3 gene copy number. The reagent kit comprises a first group of primers, i.e. target gene real-time fluorescence quantitative PCR (Polymerase Chain Reaction) amplification primers, a first fluorescent probe, i.e. a target gene real-time fluorescence quantitative PCR amplification fluorescent probe, a second group of primers, i.e. a reference gene real-time fluorescence quantitative PCR amplification primers, a second fluorescent probe, i.e. a target gene real-time fluorescence quantitative PCR amplification fluorescent probe, a calibration sample DNA (Deoxyribose Nucleic Acid), and real-time fluorescence quantitative PCR reaction liquid, wherein base sequences of the first group of primers is shown by SEQ (Sequence) ID (Identity) No. 1 and 2; a middle probe base sequence is shown by SEQ ID No. 3; base sequences of the second group of primers are shown by SEQ ID No. 4 and 5; a middle probe base sequence is shown by SEQ ID No. 6. The reagent kit for detecting the human alpha defensin 1/3 gene copy number has the advantages of simplicity, convenience and quickness in use, no standard curve required, high sensitivity and specificity, good repeatability and high detection efficiency and can be used for indicating the risk prediction of clinical infection immune diseases.

Description

technical field [0001] The invention relates to the technical field of biological diagnosis, in particular to a kit and a method for detecting the copy number of human alpha defensin 1 / 3 gene. Background technique [0002] Defensin is an endogenous cationic antibacterial peptide with a relative molecular mass of 4-5KD, containing six cysteine ​​residues and three pairs of disulfide bonds. According to the arrangement of disulfide bonds in polypeptides, defensins are divided into two categories: α-defensins and β-defensins. The gene encoding α-defensin is located in the region of chromosome 8p22-23, and there is a copy number polymorphism (CNP; copy number variation, CNV). α-defensins are mainly expressed in neutrophils, and have biological effects such as promoting the recruitment of neutrophils, enhancing the phagocytosis of macrophages, regulating complement activity, chemoattracting dendritic cells and T cells, and playing a role in the body's natural immune response an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 方向明舒强程宝莉陈齐兴金悦
Owner ZHEJIANG UNIV
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