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Application of Hsa-miR-3200-5P in preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumours

A kit and positive technology, applied in the field of medical biological detection, to achieve the effect of saving medical costs, reducing mortality, and preventing metastasis

Inactive Publication Date: 2015-09-02
SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The current research has not seen microRNAs associated with tumor-specific targets as markers for early diagnosis of tumors (including lung cancer, etc.), and there is no kit for detecting the content of this series of microRNAs in serum to target the overexpression of some Brachyury Screening of patients with tumors (including lung cancer, etc.)

Method used

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  • Application of Hsa-miR-3200-5P in preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumours
  • Application of Hsa-miR-3200-5P in preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumours
  • Application of Hsa-miR-3200-5P in preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumours

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Experimental program
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Effect test

Embodiment 1

[0046]Embodiment 1: the preparation of kit of the present invention

[0047] In order to find the microRNAs regulated by the tumor-specific transcription factor Brachyury, the inventors used lentivirus infection to transfer the cds sequence of exogenous Brachyury in a lung cancer cell line H1299 and 293T as a normal human control cell to overexpress it Then microRNA chip technology was used to screen out several microRNAs that were significantly up-regulated after being regulated by Brachyury (see figure 1 , figure 2 ).

[0048] The expression changes of these microRNAs were verified by Real-Time PCR to be consistent with the chip results. Subsequently, in the clinical plasma collection and detection of lung cancer patients, chronic obstructive pneumonia patients, and normal people, it was found that hsa-miR-3200-5p in the microRNAs group was significantly highly expressed in lung cancer patients, while in other lung parenchyma There was no significant abnormality in the ...

Embodiment 2

[0049] Embodiment 2: the preparation of kit of the present invention

[0050] The kit of the present invention consists of:

[0051] A. 2 tubes of specific reverse transcription primers, 10uM concentration, 200uL;

[0052] B. Real-Time PCR amplification specific upstream primer 1 tube (A) 10uM concentration, 200uL / tube; Real-Time PCR amplification downstream primer 1 tube 10uM concentration 2mL / tube;

[0053] C. 1 tube of standard containing quantitative microRNAs; (this kit does not contain external reference genes)

[0054] This group of microRNAs-specific reverse transcription primers is based on the mature microRNAs sequence to add a fixed Loop (loop structure) fragment, and then synthesized by the American Invitrogen Company, the purity is PAGE grade, and the synthesized primers are DEPC H 2 O was dissolved to a final concentration of 10 μM.

[0055] Specific microRNAs upstream and downstream primers: add empirical sequences based on the specific sequences of mature mi...

Embodiment 3

[0057] Embodiment 3: the detection method of this kit

[0058] (1), collecting plasma samples, extracting microRNAs and reverse transcription:

[0059] Collecting peripheral blood for detection has the advantages of non-invasive and convenient collection. Therefore, early screening of lung cancer by collecting and detecting a series of microRNAs molecular markers in peripheral blood is not only easy to popularize, but also has the advantages of high accuracy. Timely detection of early Brachyury-positive lung cancer can achieve the goal of early detection and early treatment, and improve the survival rate of patients.

[0060] Plasma samples for verification were collected from the inpatient department and outpatient department of Shanghai Changzheng Hospital, 30 specimens of primary lung cancer or bone metastases of lung cancer (selected 10 patients whose expression of Brachyury had been verified by immunohistochemistry), 30 healthy volunteer specimens ( Randomly select 10 of...

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Abstract

The invention relates to the technical field of medical biological detection. According to the invention, the microRNA chip technology is adopted to screen a plurality of microRNAs remarkably up-regulated after being regulated and controlled by Brachyury and Real-Time PCR verification is carried out on the series of microRNAs. The invention provides a molecular marker of the Brachyury positive tumours, and the molecular marker miR-3200-5p presents remarkable high expression in patients with lung cancer and presents no abnormal expression level in patients with other lung solid lesions and normal people. The invention also provides application of Hsa-miR-3200-5P in preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumours. The miR-3200-5P provided by the invention can screen and diagnose tumours in the early period of tumours, and has significant benefits for the early treatment of tumours and the reduction of medical cost.

Description

technical field [0001] The invention relates to the technical field of medical biological detection, and relates to the application of microRNAs in the preparation of reagents or kits for early screening or diagnosis of Brachyury positive tumors. Background technique [0002] Lung cancer is currently the malignant tumor with the highest mortality rate in the world. About 170,000 people die of lung cancer in the United States every year, accounting for 30% of the overall cancer deaths. With the rapid economic development and increasing air pollution, the incidence of lung cancer in my country is also showing a rising trend year by year, and it has become the leading cause of cancer death in my country's urban population. According to statistics, the five-year survival rate of lung cancer patients is only 5-15%. The main reason is that it is difficult to detect early, and most patients have metastases to varying degrees when they see a doctor. For example, in the autopsy of ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12Q1/68
Inventor 肖建如陈素矫健周振华姜东杰
Owner SECOND MILITARY MEDICAL UNIV OF THE PEOPLES LIBERATION ARMY
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