Kit for serum/plasma lncRNA marker related to stomach cancer

A kit and marker technology, applied in the fields of genetic engineering and oncology, can solve the problems of low sensitivity and specificity of the kit, insufficient specificity, increased misdiagnosis rate and missed diagnosis rate of gastric cancer, and achieve rapid and accurate control Improve the patient's condition, improve the sensitivity and specificity, and facilitate the diagnosis

Inactive Publication Date: 2016-05-18
张国新 +1
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  • Description
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AI Technical Summary

Problems solved by technology

However, the expression level of serum / plasma lncRNA is still relatively low. Without the standardization steps of standardized kits, serum / plasma lncRNA is not easy to detect, and the possibility of early diagnosis is lost; on the other hand, only one serum / plasma lncRNA is used as Tumor markers are often not specific enough, because only one serum / plasma lncRNA is used as a tumor marker kit with low sensitivity and specificity, which leads to a great increase in the misdiagnosis rate and missed diagnosis rate of early gastric cancer diagnosis
Without relatively stable biomarkers for serological diagnosis of early gastric cancer patients, it is impossible to screen abnormally expressed serum / plasma lncRNAs as biomarkers and develop corresponding diagnostic kits

Method used

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  • Kit for serum/plasma lncRNA marker related to stomach cancer
  • Kit for serum/plasma lncRNA marker related to stomach cancer
  • Kit for serum/plasma lncRNA marker related to stomach cancer

Examples

Experimental program
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Effect test

Embodiment 1

[0063] The collection of embodiment 1 sample and the arrangement of sample data

[0064] The cases were new cases of gastric cancer collected in Jiangsu Provincial People's Hospital and Jingjiang People's Hospital from June 2010 to December 2014, all of which were confirmed by histopathology. Controls were healthy individuals who underwent community disease screening during the same period, and were frequency-matched with cases by sex and age. The samples used for the research were collected at the same period, and the sampling, sub-packaging, and storage conditions were uniform. Through the arrangement of the sample data, the inventor selected 6 samples that met the following criteria as the lncRNA chip detection and subsequent series of qRT-PCR verification. Experimental sample:

[0065] 1. New gastric cancer cases

[0066] 2. No surgery, radiotherapy and chemotherapy before blood collection, no preoperative radiotherapy and chemotherapy

[0067] 3. Healthy controls match...

Embodiment 2

[0069] Example 2 lncRNA chip detection of lncRNA in serum / plasma

[0070] 3 cases of gastric cancer patients and 3 cases of healthy controls were detected by lncRNA chip to obtain relevant results. The specific steps are:

[0071] 1. Take 1ml of serum from patients in the “stomach cancer case” group and the “healthy control” group, add 1ml of Trizol reagent of equal volume, and place on ice for 5 minutes;

[0072] 2. Add 1ml of chloroform / tube and shake vigorously for 15s. Incubate at 15-30°C for 2-3min, and centrifuge (4°C, 12,000g, 15min). After centrifugation, the liquid is divided into three layers (the upper layer-colorless water sample layer is RNA, the middle white layer is DNA and the bottom red layer is protein), carefully pipette the upper layer colorless liquid into a new EP tube.

[0073] 3. Add an equal volume of isopropanol, 0.4-0.5ml, mix well, incubate at 15-30°C for 10-30min, and centrifuge (4°C, 12,000g, 10min). Remove the supernatant, add 1ml of 75% etha...

Embodiment 3

[0079] qRT-PCR experiment of lncRNA in embodiment 3 serum / plasma

[0080] According to the above chip results, lncRNAs that meet the following conditions were selected for further verification by qRT-PCR:

[0081] 1) The CT values ​​of the two groups of research objects in the chip are not greater than 35 to improve the detection efficiency;

[0082] 2) ΔΔC in the chip T greater than 2. Primers for qRT-PCR were designed for the selected two lncRNAs of MALAT1 and HOTAIR (see Table 1). The qRT-PCR detection of lncRNA was performed on individual individuals in the serum of the "gastric cancer case" group and the "healthy control" group. Strict quality control was implemented throughout the study. Each sample was tested three times consecutively. All assays were blinded, that is, performed without knowledge of the sample background to avoid bias. qRT-PCR detection was performed using the SYBR dye method.

[0083] Table 1 Related lncRNA primer information

[0084]

[0085]...

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Abstract

The invention relates to a kit for a serum/plasma lncRNA marker related to stomach cancer. The serum/plasma lncRNA marker is combination of MALAT1 and HOTAIR, a primer is a primer for the MALAT1 and the HOTAIR, and the upstream primer sequence and the downstream primer sequence of the MALAT1 are SEQ No.1 and SEQ No.2; the upstream primer and the downstream primer of the HOTAIR are SEQ No.3 and SEQ No.4. By means of the kit, the defect that a kit only adopting one serum/plasma lncRNA as the tumor marker is low in sensitivity and specificity, so that the fault diagnosis rate and the rate of early gastric cancer diagnosis are greatly raised is overcome; by means of the kit, the sensitivity and specificity of disease diagnosis are enhanced, successful development of the lncRNA biological marker is beneficial to auxiliary diagnosis of stomach cancer and provides reference for research of biological markers of other diseases, lncRNA chip detection is adopted to obtain a serum/plasma lncRNA expression profile for disease specificity and abnormity expression, a qRT-PCR method is applied to conduct single verification, and dye method is adopted to conduct verification.

Description

technical field [0001] The invention belongs to the fields of genetic engineering and oncology, and relates to a serum / plasma lncRNA marker kit related to gastric cancer. Background technique [0002] Gastric cancer is the second most common tumor in the world, and it is also one of the most common tumors in the gastrointestinal tract. Although the degree of malignancy of gastric cancer is relatively high, if early detection and early treatment can still achieve the best therapeutic effect, the surgical resection rate of early gastric cancer is 90%-100%, and the 5-year survival rate can reach 70%-100%. Compared with gastric cancer, there is a huge contrast in the treatment effects of the two. However, most patients are already at an advanced stage when they see a doctor. At present, a large number of extensive studies have been conducted on the biological markers of gastric cancer, including gene markers, serological markers, and cytological markers. Due to the differences...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6886C12Q2600/158C12Q2600/178
Inventor 张国新周晓颖
Owner 张国新
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