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Long noncoding RNA (LncRNA), and primer pair and kit for detecting expression level of long noncoding RNA in cells and tissues

A long-chain non-coding and expression-level technology, applied in the field of primers and kits for the expression level of the long-chain non-coding RNA, can solve the problem that the clinical value of the lncRNA mechanism of action needs to be further studied, and achieve far-reaching clinical significance and popularization. High sensitivity and good stability

Active Publication Date: 2015-09-02
GENERAL HOSPITAL OF PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there is still little research on lncRNAs related to the occurrence and development of gastric cancer, and the mechanism and clinical value of lncRNAs in gastric cancer still need to be further studied

Method used

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  • Long noncoding RNA (LncRNA), and primer pair and kit for detecting expression level of long noncoding RNA in cells and tissues
  • Long noncoding RNA (LncRNA), and primer pair and kit for detecting expression level of long noncoding RNA in cells and tissues
  • Long noncoding RNA (LncRNA), and primer pair and kit for detecting expression level of long noncoding RNA in cells and tissues

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. Template preparation (total RNA extraction and reverse transcription process)

[0031] Preparation of RNA: Gastric cancer tissue samples and corresponding paracancerous tissue samples as well as colorectal cancer tissue samples and corresponding paracancerous tissue samples were obtained. The paired paracancerous tissue samples GN1-GN5, and in this embodiment, 5 colorectal cancer tissue samples CRC1-CRC5, and 5 paracancerous tissue samples CN1-CN5 paired with colorectal cancer tissues.

[0032] The method of TRIZOL extraction was used to extract total RNA from the above samples respectively, the absorbance (A) value was measured by ultraviolet spectrophotometer to determine its content and purity, and the quality of RNA was identified by 1% agarose gel electrophoresis.

[0033] cDNA synthesis: 5 μg of total RNA extracted from the above-mentioned gastric cancer tissue and corresponding para-cancerous tissues and colorectal cancer tissue and corresponding para-cancerou...

Embodiment 2

[0069] Example 2: Clinical Specimen Detection

[0070] (1) Take 35 pairs of matched gastric cancer tissues and paracancerous tissues, and perform semi-quantitative RT-PCR amplification of Lnc21q22.11. Template preparation, PCR amplification system and conditions, and detection of amplification products are all the same as those in Implementation 1. , the detection results (the expression level is the relative level of the matched gastric cancer tissue and the adjacent tissue) please see the following table 1:

[0071] Table 1

[0072]

[0073] (2) Take 10 pairs of matched colorectal cancer tissues and para-cancerous tissues, and carry out semi-quantitative RT-PCR amplification of Lnc21q22.11. Template preparation, PCR amplification system and conditions, and detection of amplification products are all the same as those in the implementation The detection results (the expression level is the relative level between the matched colorectal cancer tissue and the adjacent tissue...

Embodiment 3

[0076] Embodiment 3: Sensitivity experiment

[0077] The cDNA of gastric cancer cell line BGC823 with high expression of Lnc21q22.11 was combined with ddH 2 O is mixed in proportion, and the above-mentioned Lnc21q22.11 semi-quantitative RT-PCR primer pair is used to detect the expression level of Lnc21q22.11, and the amplified band is as follows Figure 5 shown. The cDNA content of BGC823 cells was 100%, 50%, 5%, 1%, 0.5%, 0%. wxya 2 O is the system control, used to evaluate whether there is contamination of PCR products in the system, such as ddH 2 If the result of O test is negative, the result of the system is credible, and the size of the amplification product of this system is 587bp.

[0078] Figure 5 The results show that: using the Lnc21q22.11-specific primer pair, reaction system and conditions of the present invention to detect the expression of Lnc21q22.11 in cells, the sensitivity can reach 1%, and the sensitivity is high.

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Abstract

The invention provides a new long noncoding RNA (LncRNA) Lnc21q22.11 sequence, and a primer pair and kit for detecting expression level of the long noncoding RNA in cells and tissues. The full length of the LncRNA Lnc21q22.11 is identified to be 1202bp for the first time, the specific primers are utilized for the first time to detect the expression level of the LncRNA Lnc21q22.11 in stomach cancer cells, 35 pairs of stomach cancer line and para-carcinoma tissues, colorectal cancer cell lines and 10 pairs of colorectal cancer and para-carcinoma tissues, and thus, the kit is creative. The detection of the expression of the LncRNA Lnc21q22.11 in the stomach cancer tissues by using the kit and specific primers can be used as an effective means for stomach cancer and colorectal cancer diagnosis, curative effect observation, prognosis judgment, focus residue and relapse detection and the like. The kit is simple to operate, has the advantages of high stability and high sensitivity, and has far-reaching clinical meanings and popularization performance.

Description

technical field [0001] The invention relates to a novel long-chain non-coding RNA and primers and kits for detecting the expression level of the long-chain non-coding RNA in cells and tissues. Background technique [0002] According to the 2012 global cancer epidemiological statistics (GLOBOCAN2012), there were about 1 million new cases of gastric cancer (952,000, 6.8%) in 2012, which is the fifth most common malignant tumor in the world; gastric cancer is the third among the world's deadliest cancers (723000, 8.8%). More than 70% of gastric cancer cases occur in developing countries, and 50% of the worldwide total occurs in East Asia, mainly in China. Gastric cancer is the second most common malignancy in China and the third leading cause of death (Ferlay J, Soerjomataram I, Ervik M, Dikshit R, Eser S, Mathers C, Rebelo M, Parkin DM, Forman D, Bray, F. [0003] GLOBOCAN 2012v1.0, Cancer Incidence and Mortality Worldwide: IARC CancerBase No.11 [Internet]. Lyon, France: Int...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 郭明洲曹宝平张美英令狐恩强杨帅金永帅
Owner GENERAL HOSPITAL OF PLA
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