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LncRNA marker for lung cancer diagnosis and treatment

A technology of lung adenocarcinoma and substances, which is applied in the field of biomedicine and can solve the problems of less applied research

Active Publication Date: 2019-07-23
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are still relatively few applications of lncRNAs in the early diagnosis and prognosis prediction of lung adenocarcinoma, and more lung cancer-related lncRNAs need to be further explored

Method used

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  • LncRNA marker for lung cancer diagnosis and treatment
  • LncRNA marker for lung cancer diagnosis and treatment
  • LncRNA marker for lung cancer diagnosis and treatment

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] Example 1 Screening for Gene Markers Related to Lung Adenocarcinoma

[0085] 1. Sample collection

[0086] A total of 35 lung adenocarcinoma tissues and corresponding paracancerous tissues were collected. All patients had not received other treatments before surgery. Among them, 4 lung adenocarcinoma tissues and corresponding paracancerous tissues were selected for high-throughput sequencing.

[0087] 2. RNA sample preparation and quantitative analysis

[0088] Tissue RNA was extracted using QIAGEN tissue RNA extraction kit, and the specific operation was carried out according to the instructions.

[0089] The above-mentioned extracted RNA was subjected to agarose gel electrophoresis, and the concentration and purity of the extracted RNA were detected by Nanodrop2000, the RNA integrity was detected by agarose gel electrophoresis, and the RIN value was determined by Agilent2100. A single library construction requires a total amount of RNA of 5 μg, a concentration ≥ 200...

Embodiment 2

[0107] Example 2 QPCR sequencing verification of differential expression of RP11-284F21.10 gene

[0108] 1. The differential expression of RP11-284F21.10 gene was verified on 35 collected samples.

[0109] 2. RNA extraction

[0110] RNA samples were extracted using the QIAGEN Tissue RNA Extraction Kit. For details, please refer to the instruction manual.

[0111] 3. QPCR

[0112] 1) Reverse transcription reaction

[0113] Use the FastQμant cDNA First Strand Synthesis Kit (Product No.: KR106) from Tiangen Company to carry out lncRNA reverse transcription, first remove the genomic DNA reaction, add 5×gDNA Bμffer 2.0μl, total RNA 1μg, add RnaseFree ddH 2 O to bring the total volume to 10 μl, and heat in a water bath at 42°C for 3 min.

[0114] 10× Fast RT Bμffer 2.0μl, RT Enzyme Mix 1.0μl, FQ-RT Primer Mix 2.0μl, RNase Free ddH 2 O 5.0 μl, after mixing, add to the above test tube and mix together a total of 20 μl, heat in a water bath at 42°C for 15min, then at 95°C for 3min...

Embodiment 3

[0134] Example 3 Silencing of the RP11-284F21.10 gene

[0135] 1. Cell culture

[0136] Human lung adenocarcinoma cell line A549 was incubated in RPMI1640 medium containing 10% fetal bovine serum and 1% P / S at 37°C, 5% CO 2 , Cultivated in an incubator with a relative humidity of 90%. Change the medium once every 2-3 days, and use 0.25% EDTA-containing trypsin for routine digestion and passage.

[0137] 2. Design of siRNA

[0138] Design siRNA for the sequence of RP11-284F21.10 gene, the designed siRNA sequence is as follows:

[0139] Sequence of negative control siRNA-NC:

[0140] Sense strand: 5'-UUCUCCGAACGUGUCACGU-3' (SEQ ID NO.5),

[0141] Antisense strand: 5'-ACGUGACACGUUCGGAGAA-3' (SEQ ID NO.6);

[0142] siRNA1:

[0143] Sense strand: 5'-AUUGUGUAGGCUCAUGUUGCU-3' (SEQ ID NO.7),

[0144] Antisense strand: 5'-CAACAUGAGCCUACACAAUAU-3' (SEQ ID NO.8);

[0145] siRNA2:

[0146] Sense strand: 5'-AUUGCAAUUACUUCUACAGGC-3' (SEQ ID NO.9),

[0147] Antisense strand: 5'-CU...

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Abstract

The invention discloses a lncRNA marker for diagnosis and treatment, and the lncRNA marker is RP11-284F21.10. The invention firstly finds that RP11-284F21.10 expression is significantly up-regulated in lung adenocarcinoma, and further proves that RP11-284F21.10 can be used as a potential molecular target for clinical diagnosis of lung adenocarcinoma by large sample QPCR experiments. The inventionsimultaneously proves that RP11-284F21.10 affects the proliferation of lung adenocarcinoma cells by CCK8 experiments.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to lncRNA markers for diagnosis and treatment of lung cancer. The specific marker is RP11-284F21.10. Background technique [0002] Lung cancer is the cancer with the highest morbidity and mortality in the world (FERLAY J, SOERJOMATARAM I, DIKSHITR, et al. Cancer incidence and mortality worldwide: sources, methods and major patterns in GLOBOCAN 2012[J]. Int J Cancer.2015, 136( 5): E359-386.). Lung cancer includes four main subtypes, and several minor or extremely rare subtypes (BRAMBILLA E, TRAVIS W D, COLBY T V, et al. The new World Health Organization classification of lung tumors[J]. EurRespir J.2001, 18(6):1059-1068.), each subtype has specific biological and clinical features. They can be divided into small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) by clinicopathological features (SUN S, SCHILLER J H, GAZDAR A F. Lung cancer never smokers--a different disease[J].N...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886A61K45/00A61P35/00
CPCA61K45/00A61P35/00C12Q1/6886C12Q2600/136C12Q2600/158C12Q2600/178
Inventor 杨承刚高舒欣
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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