Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Probes, primers and reagent kits for detecting five mutations of human gene TP53

A technology for detecting probes and kits, which is applied in the biological field and can solve problems such as easy contamination of samples, difficulty in differentiation, and poor accuracy

Active Publication Date: 2016-04-06
武汉海吉力生物科技有限公司
View PDF2 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problems of the existing TP53 gene mutation detection methods, such as low detection efficiency, easy sample contamination, long detection time, poor accuracy, and difficulty in discrimination, the present invention provides primers and probes for detecting five mutations of the human TP53 gene

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Probes, primers and reagent kits for detecting five mutations of human gene TP53
  • Probes, primers and reagent kits for detecting five mutations of human gene TP53
  • Probes, primers and reagent kits for detecting five mutations of human gene TP53

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] 1. Primers and probes of the detection kit for detecting 7 mutations of human TP53 gene:

[0079] According to the wild-type sequence of the TP53 gene (NCBIReferenceSequence: NM_000546.5) published in the NCBI database, the R175H (TM1) mutation site in exon 5, the R248W (TM2) and R248Q (TM3) mutation sites in exon 7, and the exon 8 Using exon R273C (TM4) and R273H (TM5) mutation sites as reference, design specific R175H mutation primers and probes (see Table 2), specific R248W mutation primers and probes (see Table 3), specific R248Q mutation Primers and probes (see Table 4), specific R273C mutation primers and probes (see Table 5) and specific R273H mutation primers and probes (see Table 6). Using the mutant plasmid and wild-type plasmid constructed by genetic engineering as templates, a real-time fluorescent PCR mutation detection system was established to achieve high sensitivity and high specificity detection of TP53 gene mutation.

[0080] (1) Specific primer and ...

Embodiment 2

[0156] The human TP53 gene 5 mutation detection kit of the present invention is used to detect clinically collected tissue samples.

[0157] In this example, tissue samples from patients diagnosed as breast cancer were collected and genomic DNA was extracted from them. The TP53 gene mutation detection kit was used to detect the mutation status of the TP53 gene in the samples to be tested. Specific steps are as follows:

[0158] (1) Sample genomic DNA extraction

[0159] Paraffin-embedded tissue samples were extracted using QIAampDNAFFPETissueKit (CatNo.56404). Extract the genomic DNA of the sample to be tested according to the instructions, and use the ultraviolet spectrophotometer to detect the concentration and DNA quality of the extracted DNA sample. The concentration of DNA is required to be ≥10ng / μL, and the quality of DNA OD260 / 280 is between 1.8 and 2.0. Genomic DNA that meets the requirements was diluted to 2-10 ng / μL with TE (10 mmol / L, pH 8.0) as a PCR template fo...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses probes, primers and reagent kits for detecting five mutations of a human gene TP53. According to five primers and probes for TM1-TM5, each mutation primer can be combined with a conserved sequence of the gene TP53, the primer for the mutation ARMS can be specifically combined with the corresponding mutation sequence, and the mutation sequence is amplified. The detection probe can be combined with amplified fragments, the blocking probe can be specifically combined with a wild type sequence corresponding to a mutation site, and wild type non-specificity amplification is restrained. The specificity mutation primers and the probe blocking technology are adopted, and the five mutation types of the gene TP53 can be accurately detected; mutations of the gene TP53 are rapidly detected conveniently through the built reagent kits of a real-time fluorescence PCR amplified reaction system, operation is easy, and the result is easy to read. The detection method is high in sensitivity, and 50-copy mutation can be stably detected; the specificity is good, 20 ng wild type genomic DNA non-specificity amplification is achieved, and the detectability reaches up to 1%.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a probe, primer and kit for detecting five mutations of human TP53 gene. Background technique [0002] The wild-type TP53 gene is a tumor suppressor gene, and the p53 protein encoded by it can repair damaged cells or remove severely damaged cells so as to avoid the harm of these cells to the body. TP53 gene mutations are related to 50% of human tumors, currently found are liver cancer, breast cancer, bladder cancer, gastric cancer, colon cancer, prostate cancer, soft tissue sarcoma, ovarian cancer, brain tumor, lymphocytic tumor, esophageal cancer, lung cancer, Osteogenic sarcoma, etc., and there are differences in TP53 allele mutations in different tumors. [0003] TP53 mutations in human tumors are mainly in highly conserved regions, most of which are missense mutations, among which the mutations at codon sites such as 175, 245, 248, 249, 273, and 283 are the highest, which may be...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q1/6886C12Q2600/106C12Q2600/156C12Q2531/113C12Q2535/137C12Q2537/163C12Q2561/101C12Q2561/113
Inventor 朱峰杨惠娟段卫涛赵平锋
Owner 武汉海吉力生物科技有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com