Non-small cell lung cancer marker, detection reagent and kit

A non-small cell lung cancer, detection kit technology, applied in the field of molecular biology, can solve the problems of patients losing surgically resected lesions, unsatisfactory detection sensitivity and specificity, and inconspicuous early symptoms of lung cancer. Sensitivity and specificity, easily detectable effects

Inactive Publication Date: 2016-12-07
上海晟燃生物科技有限公司
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Problems solved by technology

However, due to the limitations of such methods, the sensitivity and specificity of the detection itself are not very satisfactory
The early symptoms of lung cancer are n

Method used

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  • Non-small cell lung cancer marker, detection reagent and kit
  • Non-small cell lung cancer marker, detection reagent and kit
  • Non-small cell lung cancer marker, detection reagent and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1 Screening of miRNAs associated with human lung cancer

[0059] 1.1 Collection of samples and collation of data

[0060] The inventor collected a large number of peripheral blood samples from lung cancer patients, pneumonia patients and healthy individuals in the Affiliated Hospital of Wenzhou Medical University in November 2014 (sample collection, pretreatment, subpackaging, storage conditions, etc. of different batches are consistent) , through sorting out the sample data, the inventor selected 30 cases of plasma from patients with non-small cell lung cancer randomly collected in the same hospital and as far as possible in the same department, 20 cases of plasma from patients with pneumonia randomly collected in the same hospital and the same period as the experimental group, and 40 cases of plasma from the same hospital and the experimental group. Plasma samples from healthy individuals randomly collected from the same period as the experimental group (inclu...

Embodiment 2

[0082] Example 2 QPCR verification of differentially expressed miRNA-375

[0083] According to the detection results of the miRNA chip above, miRNA-375 was selected for QPCR verification of a larger number of samples. 95 samples from the non-small cell lung cancer group and 120 samples from the healthy group were selected according to the method of sample collection and sample data arrangement in Example 1.

[0084] 2.1 The extraction process of total RNA is the same as in Example 1.

[0085] 2.2 Reverse transcription: mix 10pg-1μg total RNA template with 2μl 5* buffer, 1μl genomic DNA digesting enzyme and RNase-free water (RNase-free water) to a final volume of 10μl, incubate at 42°C for 5min Genomic DNA is removed. Then add 0.5 μl of 20 μM specific reverse transcription primer to the reaction tube, incubate at 65°C for 10 minutes and immediately incubate on ice for at least 2 minutes to break the secondary structure of RNA and primers. Finally, mix the above reaction mixt...

Embodiment 3

[0087] Example 3 Analysis of the prediction of exosome-derived miRNA on the incidence of non-small cell lung cancer

[0088] Exosome-derived miRNA-375 was used as the research object and compared with plasma free miRNA in terms of sensitivity and specificity in distinguishing control and lung cancer groups. Taking patients with non-small cell lung cancer as the experimental group, and patients with pneumonia and normal people as the control group, firstly use the total exosome isolation reagent (Invitrogen) to extract the exosomes in the plasma (the steps are basically the same as in Example 1). 40 non-small cell lung cancer plasma samples, 20 pneumonia patients, and 40 normal human plasma samples were selected for exosomal miRNA-375 (exosome group) and plasma free miRNA-375 (plasma group) QPCR experiments (method basic Same as Example 2, but the plasma group is directly extracting free miRNA-375 instead of separating from exosomes), and the CT value of each specimen is obtain...

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Abstract

The invention discloses a blood miRNA marker related to human non-small cell lung cancer and an application of the blood miRNA marker. The marker is plasma exosome-derived miRNA-375. The invention also provides a detection reagent of the blood miRNA marker related to the human lung cancer and a kit containing the detection reagent. On the other side, the invention also provides a method for detecting the marker miRNA-375. The marker and the detection reagent thereof provided by the invention can be used for preparing the detection kit for the early prognosis and detection of the non-small cell lung cancer. Different from conventional biological markers, the exosome-derived miRNA marker can achieve a stable and minimally invasive operation, easy detection and precise quantification, and meanwhile, the sensitivity and the specificity of disease diagnosis can be greatly improved.

Description

technical field [0001] The invention belongs to the field of molecular biology, and in particular relates to a miRNA marker and its application. Background technique [0002] Lung cancer is one of the most common malignant tumors in the world. In the past 50 years, many countries have reported that the morbidity and mortality of lung cancer have increased significantly. The incidence and mortality of lung cancer in men rank first among all malignant tumors, and the incidence and mortality of lung cancer occupy the second place in women. Lung cancer has also leapt to become the first cause of death from malignant tumors in my country's urban population. Non-small cell lung cancer accounts for about 80% of all lung cancers, including squamous cell carcinoma (squamous cell carcinoma), adenocarcinoma, etc. Compared with small cell carcinoma, non-small cell lung cancer cells grow and divide more slowly, and spread and metastasize relatively later. About 75% of non-small cell l...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12Q1/68
Inventor 王家亮王昉炜李辉辉刘月星
Owner 上海晟燃生物科技有限公司
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