Circ_3234, application thereof in preparation of nasopharyngeal carcinoma therapeutic preparations and therapeutic preparation of nasopharyngeal carcinoma

A technology for nasopharyngeal carcinoma and preparations, 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 important promotion and application prospects, good silent effect, and profound clinical significance.

Active Publication Date: 2019-07-05
CENT SOUTH UNIV
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  • Abstract
  • Description
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  • Application Information

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Problems solved by technology

Therefore, the specific molecular mechanism of the occurrence and development of nasopharyngeal carcinoma is still unclear, and due to tum

Method used

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  • Circ_3234, application thereof in preparation of nasopharyngeal carcinoma therapeutic preparations and therapeutic preparation of nasopharyngeal carcinoma
  • Circ_3234, application thereof in preparation of nasopharyngeal carcinoma therapeutic preparations and therapeutic preparation of nasopharyngeal carcinoma
  • Circ_3234, application thereof in preparation of nasopharyngeal carcinoma therapeutic preparations and therapeutic preparation of nasopharyngeal carcinoma

Examples

Experimental program
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Example Embodiment

[0082] Example 1: Detection of circ_3234 overexpression effect in nasopharyngeal carcinoma cell lines

[0083] First, we select the restriction site and put the full-length sequence of circ_3234 into the NEB cutter 2.0 online website for analysis. It shows that the restriction sites of Sacll and Clal are not present in the full-length sequence of circ_3234, and they are also in the pcDNA3.1 plasmid vector ( (Purchased from Shenggong Biological Engineering Co., Ltd.) in a single DNA restriction endonuclease. According to this, the expression vector was constructed, see figure 1 .

[0084] In order to detect the efficiency of circ_3234 looping, we first expressed the constructed pcDNA3.1 / circ_3234 eukaryotic overexpression vector in nasopharyngeal carcinoma cells. Plant the third and fourth generation nasopharyngeal carcinoma cells HONE1, HNE2 and CNE2 in a good growing condition into a 12-well plate. When the cell confluence reaches 60%-80%, use lipofectamine 3000 to remove endotox...

Example Embodiment

[0085] Example 2: Detection of the effect of silencing circ_3234 expression in nasopharyngeal carcinoma cell lines

[0086] The circ_3234 siRNA sequence was designed according to the splicing site. siRNA is a type of double-stranded RNA molecule with a length of 21-25 nucleotides that can complementally bind to homologous RNA and specifically degrade the target RNA, thereby inhibiting its expression. At present, siRNA has developed into an important tool for gene function research. In order to explore the role of circ_3234 in tumorigenesis and development, we designed siRNA based on the splicing site of circ_3234, and transiently transfected siRNA and siNC (blank control) into HONE1, HNE2 and CNE2 cell lines to silence the expression of circ_3234 using Hiperfect reagent. After transfection, continue to culture for 48 hours to collect cells, and use real-time fluorescent quantitative PCR to detect the expression level of circ_3234 to detect the transfection efficiency of siRNA. It...

Example Embodiment

[0087] Example 3: Cell Scratch Healing and Migration Experiment:

[0088] (1) Cell imaging table: 1000μl / 10μl Tip, high-temperature autoclave D-Hank's, ruler, 1000μl / 10μl pipette, marker pen, etc., sterilized with alcohol and placed in the ultra-clean table after 30 minute;

[0089] (2) When the cells grow to about 50% to 70%, transfect siRNA and NC groups or transfect plasmids respectively;

[0090] (3) Scratch the next day after the cells overgrown the bottom of the flat plate: Place a 10μl pipette tip perpendicular to the bottom of the 6-well plate with a straightedge and quickly make a cross or tic-tac-toe scratch. Do not tilt, and the strength is the same. Make sure that the width of the scratch is as same as possible

[0091] (4) Aspirate and discard the culture medium, wash gently with D-hanks 3 times to wash away the debris cells caused by scratches as much as possible;

[0092] (5) 1640 medium with 1% double antibody and 2% fetal bovine serum;

[0093] (6) Take a photo and rec...

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Abstract

The invention belongs to the technical field of oncomolecularbiology and particularly relates to circ_3234, application thereof in the preparation of nasopharyngeal carcinoma therapeutic preparationsand a therapeutic preparation of nasopharyngeal carcinoma. The circ_3234 has good gene silencing effect, it is ensured that circ_3234 in nasopharyngeal carcinoma cell lines HONE1, HNE2 and CNE2 is interfered and silenced by siRNA, Transwell chamber experiment and scratch healing experiment are carried out, invasion migration speed of siRNA group cell invasion migrating speed is decreased as compared with a control group; the invasive migration of nasopharyngeal carcinoma cells is inhibited by silencing circ-3234. Correspondingly, a vector to overexpress circ_3234 is constructed; after circ_3234 is overexpressed in nasopharyngeal carcinoma lines HONE1, HNE2 and CN32, cell invasive migration speed is increased evidently; namely, circ_3234 can retard the diffusion transfer of nasopharyngeal carcinoma and has profound 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 in particular relates to a circular RNA circ_3234, a reagent for inhibiting the expression of the circular RNA circ_3234, and its application in preparing nasopharyngeal carcinoma treatment preparations. 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 formed by reverse splicing of precursor mRNA (precursor messenger RNA, pre-mRNA) without a 5' end cap and a 3' end poly(A) tail, and forms a circular structure with a covalent bond molecular. [0003] The formation process of circRNA can be divided into two categories, namely exon circularization (exon circularization) and intron cir...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12Q1/6886A61K45/00A61K31/713A61P35/00
CPCA61K31/713A61K45/00A61P35/00C12N15/113C12N2310/532C12Q1/6886C12Q2600/158C12Q2600/178
Inventor 曾朝阳郭灿熊炜范春梅赵梦瑶熊芳张立身莫勇真彭苗李征李小玲李桂源
Owner CENT SOUTH UNIV
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