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Plasma/serum circulation microRNA marker related to mlignnt melnom and application of marker

A malignant melanoma and marker technology, applied in the fields of biotechnology and medicine, to achieve the effects of improving sensitivity and specificity, accurate quantification, and easy detection

Active Publication Date: 2014-03-19
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] However, there are no reports of relatively stable circulating miRNA markers for the diagnosis of melanoma. If the circulating miRNAs abnormally expressed in melanoma can be screened out as biomarkers, and corresponding diagnostic kits can be developed, it will be beneficial to Chinese melanoma. The diagnosis status of melanoma will be a powerful impetus

Method used

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  • Plasma/serum circulation microRNA marker related to mlignnt melnom and application of marker
  • Plasma/serum circulation microRNA marker related to mlignnt melnom and application of marker
  • Plasma/serum circulation microRNA marker related to mlignnt melnom and application of marker

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Embodiment 1

[0074] The collection of embodiment 1 sample and the arrangement of sample data

[0075] The inventor has collected a large number of peripheral blood samples from patients with malignant melanoma and controls from Xijing Hospital Affiliated to the Fourth Military Medical University since 2008 (the samples used for research were collected at the same period, and the sampling, packaging, and storage conditions were uniform). For the arrangement of sample data, the inventor selected 260 samples that meet the following criteria as experimental samples for Agilent microRNA chip detection and subsequent series of qRT-PCR verification:

[0076] 1. New cases of malignant melanoma

[0077] 2. No surgery, radiotherapy and chemotherapy before blood collection, no preoperative radiotherapy and chemotherapy

[0078] 3. Healthy controls matched with the age of the cases and systematically collected demographic data, clinical data, etc. of these samples.

Embodiment 2

[0079] Agilent microRNA chip detection of miRNA in embodiment 2 serum / plasma

[0080] Among the above-mentioned eligible 30 malignant melanoma patients and 30 healthy controls, the two groups were age-matched. The two groups of people were detected by Agilent microRNA chips to obtain relevant results. The specific steps are:

[0081] 1. Use mirVana PARIS microRNA Isolation kit (mirVana PARIS miRNA Isolation kit) to extract total RNA according to the instructions provided by the manufacturer (Ambion, Austin, TX). During the extraction process, add a final concentration of 10 -4 pmol / μl artificially synthesized cel-39 (TAKARA) was used as an internal reference, and its concentration was quantitatively detected with a NanoDrop1000 spectrophotometer (NanoDrop Technologies, Waltham, MA).

[0082] 2. Label and hybridize the RNA sample. The reagents required for labeling and hybridization are included in Agilent's miRNA Complete Labeling and Hyb Kit (p / n5190-0456). The specific rea...

Embodiment 3

[0095] qRT-PCR experiment of miRNA in embodiment 3 serum / plasma

[0096] According to the above Agilent microRNA results, the top 10 miRNAs with differential expression were selected and verified by qRT-PCR in 30 cases of malignant melanoma patients and 30 cases of healthy controls:

[0097] For selected hsa-miR-610, hsa-miR-1224-5p, hsa-miR-516a-5p, hsa-miR-125a-3p, hsa-miR-202, hsa-miR-557, hsa-miR- 1182, hsa-miR-1299, hsa-miR-877, hsa-miR-371-5p and other 10 miRNAs designed qRT-PCR primers. The qRT-PCR detection of miRNA was performed on the serum individual individuals of the "malignant melanoma cases" 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. The dye method and the probe method were used for qRT-PCR detection respectively.

[0098] (1) Preparation of RNA sample...

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Abstract

The invention relates to a plasma / serum circulation microRNA marker related to mlignnt melnom and application of the marker. The plasma / serum circulation microRNA marker related to mlignnt melnom is combination of miR-610, miR-1224-5p, miR-125a-3p and miR-557. The invention provides the plasma / serum circulation microRNA marker which has the capability of early diagnosis and mlignnt melnom screening and is related to mlignnt melnom, and the application of the marker.

Description

technical field [0001] The invention belongs to the fields of biotechnology and medicine, and relates to a plasma / serum circulating microRNA marker related to malignant melanoma and an application thereof. Background technique [0002] Melanoma is a highly aggressive skin malignancy [Clark WH Jr et al, 1984], the prevalence rate is about 0.02-0.5‰ in the world, and the annual growth rate is 3-5%, which is the highest incidence rate among all malignant tumors. One of the fastest growing malignant tumors. Melanoma has a high mortality rate. Although its incidence accounts for only 4% of skin tumors, it accounts for 74% of the deaths caused by skin tumors. It has become one of the diseases that seriously endanger human health. Melanoma diagnosed and resected early has a good prognosis, with a 5-year survival rate of over 90%. However, the 5-year survival rate of patients with stage II melanoma rapidly drops to 60%, and the 5-year survival rate of patients with stage III melano...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/113
CPCC12Q1/6886C12Q2600/158C12Q2600/178
Inventor 李春英高天文郭森石琼李凯王刚
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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