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Long and non-coding RNA and application thereof in diagnosis/treatment of bile duct carcinoma

A cholangiocarcinoma, non-coding technology, applied in the field of genetic engineering, can solve the problems of lncRNA that have yet to be discovered

Inactive Publication Date: 2017-08-04
THE SECOND AFFILIATED HOSPITAL OF NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the lack of research on lncRNAs, the specific functional mechanisms of existing lncRNAs have yet to be explored, and new lncRNAs have yet to be discovered

Method used

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  • Long and non-coding RNA and application thereof in diagnosis/treatment of bile duct carcinoma
  • Long and non-coding RNA and application thereof in diagnosis/treatment of bile duct carcinoma
  • Long and non-coding RNA and application thereof in diagnosis/treatment of bile duct carcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Isolation and Identification of Exosomes

[0049] (1) Take out the patient’s bile specimen stored in the -80°C refrigerator, balance with pre-cooled PBS, centrifuge at 300xg for 10 minutes at 4°C, discard the pellet to remove the precipitated cells and cell debris, and transfer the supernatant to a new Labeled 10ml centrifuge tube.

[0050] (2) The supernatant taken out was balanced with pre-cooled PBS and then centrifuged at 16,500 x g for 20 minutes at 4°C. The precipitate was discarded to further remove the debris in the supernatant. The supernatant was sucked with a syringe and filtered through a 0.22 μm filter, and the filtrate was collected in Labeled ultracentrifuge tubes.

[0051] (3) Add precooled PBS to more than 2 / 3 of the volume of the centrifuge tube in the ultracentrifuge tube containing the filtrate and balance with PBS. Vacuum centrifuge at 120,000 x g in an ultracentrifuge for 70 minutes at 4°C, and a yellow precipitate can be observed at th...

Embodiment 2

[0053] Example 2 Detection of the expression of ENST00000588480.1R in bile exosomes

[0054] RNA extraction

[0055] After the exosomes were blown evenly with trizol and allowed to stand on ice for 5 minutes, about 0.2 ml of chloroform (trichloromethane, volume ratio of chloroform: trizol=1:5) was added, mixed quickly and vigorously, and left to stand on ice for 5 minutes;

[0056] The above mixture was centrifuged at 12000rpm / min at 4°C for 20min, and then divided into three layers (aqueous phase / white precipitate / red organic matter). Use a pipette to gently absorb the colorless and transparent liquid in the upper layer into a new enzyme-free EP tube. Avoid white precipitate;

[0057] Add an equal volume of isopropanol to the colorless and transparent supernatant, vortex at low speed, let stand on ice for 10 minutes, centrifuge at 12,000 rpm / min at 4°C for 15 minutes, and discard the supernatant;

[0058] After adding 1ml of pre-cooled freshly prepared 75% ethanol (0.1% DEP...

Embodiment 3

[0081] Example 3 The Value of ENST00000588480.1 in the Diagnosis and Prognosis Evaluation of Cholangiocarcinoma

[0082] In order to detect the value of ENST00000588480.1 in the diagnosis of cholangiocarcinoma, the ROC curve showed that the area under the curve was 0.688 (95% confidence interval: 0.55-0.82), sensitivity: 63%, specificity: 75% ( figure 2 B). High expression of ENST00000588480.1 in cholangiocarcinoma was associated with TNM staging of patients (Ⅰ-Ⅱvs.Ⅲ-Ⅳ, p=0.027, image 3 A) Correlation, according to ENST00000588480.1 in cholangiocarcinoma patients and poor prognosis (p=0.0231, image 3 B) There is a positive correlation.

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Abstract

The invention belongs to the field of genetic engineering and in particular relates to an application of a bile exosome long and non-coding RNA (lncRNA) ENST00000588480.1 in diagnosis of bile duct carcinoma, prognostic prediction and target spot drug treatment, wherein the long and non-coding RNA is positively correlated with TNM staging and poor prognosis of a patient.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and particularly relates to the application of a long non-coding RNA-ENST00000588480.1 in diagnosis and preparation of cholangiocarcinoma medicine. Background technique [0002] Cholangiocarcinoma (CCA) is a malignant tumor originating from bile duct epithelial cells, with insidious onset and difficult early diagnosis. The average survival period is less than 24 months, and the five-year survival rate is less than 5%. Surgery is currently an effective treatment, but only 30% of newly diagnosed patients are suitable for surgery, and the rest have lost their chance of surgery. Therefore, early diagnosis and early treatment are the key to improving the operation rate and survival rate of patients with cholangiocarcinoma. At present, the gold standard for the diagnosis of cholangiocarcinoma relies on histology and cytology. Endoscopic retrograde cholangiopancreatography (ERCP) is commonly used in ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12Q1/68C12N15/11A61K31/7105A61P35/00
CPCA61K31/7105C12N15/1135C12N2320/30C12Q1/6886C12Q2600/112C12Q2600/118C12Q2600/158C12Q2600/178
Inventor 缪林葛贤秀季杰李全朋聂俊杰唐玲玉
Owner THE SECOND AFFILIATED HOSPITAL OF NANJING MEDICAL UNIV
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