Application of reagent for detecting circ_CLASP2 to preparation of nasopharynx cancer diagnosis preparation as well as kit

A nasopharyngeal carcinoma and kit technology, applied in the field of tumor molecular biology, can solve the problems of unclear molecular mechanism and insignificant curative effect, and achieve the effect of true and reliable results, rigorous test process and strict screening standards

Active Publication Date: 2018-10-16
CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the specific molecular mechanism of the occurrence and development of nasopharyngeal carcinoma is still unclear, and due to tumor heterogeneity and individual differences, the curative effect of traditional radiotherapy combined with chemotherapy is not significant

Method used

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  • Application of reagent for detecting circ_CLASP2 to preparation of nasopharynx cancer diagnosis preparation as well as kit
  • Application of reagent for detecting circ_CLASP2 to preparation of nasopharynx cancer diagnosis preparation as well as kit
  • Application of reagent for detecting circ_CLASP2 to preparation of nasopharynx cancer diagnosis preparation as well as kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0096] Example 1: Expression of circ_CLASP2 in nasopharyngeal carcinoma tissues and cells

[0097] 1. According to the standard sample collection plan, we collected 34 tissue samples from patients with suspected nasopharyngeal carcinoma from Hunan Cancer Hospital. All the cases were newly diagnosed patients in the Department of Head and Neck Surgery of Hunan Cancer Hospital (time interval: January 2016 to November 2016). 24 cases of nasopharyngeal carcinoma and 12 cases of nasopharyngeal inflammation were diagnosed by the pathology department (neoplastic diseases, no active infectious diseases, severe immune diseases and other major diseases have been excluded).

[0098] Complete personal information and clinical data were recorded during the collection process, including name, gender, age, outpatient number, hospitalization number, pathological type, case stage, and EBV infection, etc. See the screenshot of the Excel spreadsheet for details. All samples are collected with th...

Embodiment 2

[0135]Example 2 Detection of circ_CLASP2 overexpression effect in nasopharyngeal carcinoma cell lines

[0136] First, we select the enzyme cutting site, and put the full-length sequence of circ_CLASP2 (that is, the 449bp sequence of exons 2 to 6 of the CLASP2 gene) into the NEB cutter 2.0 online website for analysis, showing that the ClaI and SacII enzyme cutting sites are in the full-length sequence of circ_CLASP2 A site that does not exist, and a DNA restriction enzyme that is uniquely present in the pcDNA3.1 plasmid vector. Construct an overexpression vector accordingly; image 3 Map of the overexpression vector.

[0137] In order to test the overexpression efficiency of circ_CLASP2, we first expressed the constructed circ_CLASP2 eukaryotic overexpression vector in nasopharyngeal carcinoma cells. Nasopharyngeal carcinoma cells CNE2 and HNE2 in good growth condition were seeded into 12-well plates, and when the cell confluence reached 60%-80%, the endotoxin-free plasmid bl...

Embodiment 3

[0138] Example 3 Detection of the effect of silencing circ_CLASP2 expression in nasopharyngeal carcinoma cell lines

[0139] In order to ensure the rigorousness of the experiment, we designed two siRNA sequences of circ_CLASP2 according to the splicing site. siRNA is a type of 21-25 nucleotides in length, which can complement and bind with homologous RNA and specifically degrade the target RNA, thereby Double-stranded RNA molecules that inhibit their expression. At present, siRNA has developed into an important tool for the study of gene function. In order to explore the role of circ_CLASP2 in the development of tumors, we designed two siRNAs according to the splicing site of circ_CLASP2, and used Hiperfect reagent to combine siRNAs (siRNA1, siRNA2 and their mixture siRNA1+2) and an unrelated sequence as an experimental control (NC) Transient transfection into CNE2 and HNE2 cell lines silenced the expression of circ_CLASP2. Continue to culture for 48 hours after transfection...

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Abstract

The invention discloses application of a reagent for detecting circ_CLASP2 to preparation of a nasopharynx cancer diagnosis preparation as well as a kit. A plurality of nasopharynx cancer tissues andnormal nasopharynx epithelial tissues are collected, and the expression condition of circ_CLASP2 is detected by utilizing a qRT-PCR technology. The result shows that the expression level of the circ_CLASP2 in the nasopharynx cancer tissue is three times that of the normal inflammatory nasopharynx epithelium, and the difference of the two groups of data has statistical significance (P is equal to 0.0386). Therefore, the circ_CLASP2 can serve as a nasopharynx cancer auxiliary diagnosis molecular marker and has a far-reaching clinical significance and an important popularization and application prospect.

Description

technical field [0001] The invention belongs to the technical field of tumor molecular biology, and specifically relates to the application of a reagent for detecting circular RNA circ_CLASP2 in the preparation of nasopharyngeal carcinoma diagnostic preparations and a corresponding kit. Background technique [0002] At present, circular RNA (circular RNA) is a research hotspot. CircRNA is mainly derived from the exon region of protein-coding genes, and can also be composed of intron regions, UTR regions, intergenic regions, non-coding RNA sites and known transcripts. antisense site formation. CircRNA is a kind of non-coding RNA molecule formed by reverse splicing of precursor RNA (precursor RNA, pre-RNA) without a 5' end cap and a 3' end poly(A) tail, and forms a circular structure with a covalent bond. . [0003] The formation process of circRNA can be divided into two categories, namely exon circularization (exon circularization) and intron circularization (intron circul...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/158C12Q2600/178
Inventor 曾朝阳熊炜郭灿李桂源李小玲熊芳莫勇真王裕民陶文静张姗姗龚朝建刘凌云
Owner CENT SOUTH UNIV
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