Application of set of miRNAs in preparation of marker for screening early-stage breast cancers

A breast cancer and marker technology, applied in the field of miRNA, can solve the problem of difficult to detect miRNA in early breast cancer

Pending Publication Date: 2020-05-15
北京粒基生物科技有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The present invention aims at the technical problem existing in the prior art that conventional plasma is difficult to detect miRNA in early breast can

Method used

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  • Application of set of miRNAs in preparation of marker for screening early-stage breast cancers
  • Application of set of miRNAs in preparation of marker for screening early-stage breast cancers
  • Application of set of miRNAs in preparation of marker for screening early-stage breast cancers

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] 1. Determination and collection of samples

[0056] The samples for the previous study were from 10 breastfeeding mothers, who had no past history of cancer or other major diseases, and the informed consent of the donors had been obtained for the experimental use and sample collection. The collection of breast milk is that the donor expresses himself with breast pumping equipment and transfers it to the inventor within 2 hours.

[0057] 2. Extraction and detection of total free RNA (cfRNA) in breast milk

[0058]The breast milk samples were centrifuged at 1500g for 10 minutes. After centrifugation, the milk fat was discarded, and the supernatant was transferred to a new centrifuge tube. The cell pellet at the bottom of the tube was directly used to extract total RNA. The extraction method used SUMZol RNA extraction reagent (SummerBio). The isolated supernatant was then centrifuged at 10,000g for 10 minutes to remove the top layer of milk fat, and then human milk cfRNA ...

Embodiment 2

[0083] Application of reverse transcription-fluorescence quantitative PCR method to detect miRNA as early breast cancer screening markers

[0084] 1. Extraction of breast milk free RNA (cfRNA)

[0085] The free DNA extraction kit (SummerBioSUM66001) was used to extract cfRNA from breast milk, and the operation steps were carried out according to the kit instructions. The extracted cfRNA was stored at -20°C after concentration detection.

[0086] 2. cDNA first-strand synthesis

[0087] The first strand of miRNA was synthesized by the PolyA method, and the reaction volume was 20 μl. The commercialized SUMi miRNA cDNA first-strand synthesis kit (PolyA tail method) (SummerBioSUM0201) was used in the experiment, and the specific reaction system was prepared as follows.

[0088] Table 5 Reaction system of PloyA method PCR

[0089]

[0090] Gently mix the prepared reaction solution with a pipette, centrifuge briefly, and perform the reverse transcription reaction of miRNA in t...

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Abstract

The invention discloses application of a set of miRNAs in preparation of a marker for screening early-stage breast cancers. The invention discovers that free miRNAs with completeness and relative abundance quite accordant with mammary gland cells exist in human breast milk for the first time. Through comparison with abnormal miRNAs which are abnormally expressed by the breast cancers and are confirmed by existing researches, a set of breast milk free miRNAs suitable for potential application of the screening of the early-stage breast cancers are analyzed, 26 miRNAs are expected to be regulatedup in the development process of the early-stage breast cancers, and 12 miRNAs are expected to be regulated down in the development process of the early-stage breast cancers. The invention disclosesa theoretically feasible application detection scheme. By taking breast milk as a sample, the affection risk of the early-stage breast cancers is detected by detecting free miRNA markers related withthe breast cancers in the breast milk.

Description

technical field [0001] The invention relates to miRNA, in particular to a group of breast milk free miRNA which can be potentially applied to early breast cancer screening and its application. Background technique [0002] Breast cancer is the most common cancer in women. According to the latest national breast cancer report released by the National Cancer Center in 2018, the number of female breast cancer cases and deaths in China accounted for 11.2% and 9.2% of the global incidence and death respectively, ranking among the top in the world. The incidence and death of female breast cancer rank first and fifth in the incidence and death of female malignant tumors in my country, respectively. [0003] The 5-year survival rate of breast cancer is relatively high, about 80%; the cure rate of early breast cancer can be as high as 90%. Therefore, for breast cancer, early detection and early treatment are the key. At present, the commonly used B-ultrasound of the breast under t...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/113
CPCC12Q1/6886C12Q2600/178C12Q2600/158
Inventor 宋庆贺张莹莹
Owner 北京粒基生物科技有限公司
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