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Long-chain non-coding RNA for early diagnosis of human prostate cancer and its preparation and use

A long-chain non-coding, prostate cancer technology, applied in the direction of DNA preparation, DNA / RNA fragments, recombinant DNA technology, etc., can solve the problems of low correlation and low diagnostic accuracy of prostate cancer, and achieve high correlation and step-by-step Simple, Accuracy-Enhancing Effects

Active Publication Date: 2019-06-04
SUZHOU INST OF BIOMEDICAL ENG & TECH CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] Therefore, the technical problem to be solved by the present invention is that the correlation of the long non-coding RNA prostate cancer used for early diagnosis of human prostate cancer in the prior art is not high, and the diagnostic accuracy is low, and then a method for early diagnosis of human prostate cancer is provided. The long-chain non-coding RNA of prostate cancer and its preparation and application, such as the RNA interference target site based on the long-chain non-coding RNA, the short hairpin fragment of RNAi, and the shRNA recombinant plasmid can be used for the related treatment of human prostate cancer

Method used

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  • Long-chain non-coding RNA for early diagnosis of human prostate cancer and its preparation and use
  • Long-chain non-coding RNA for early diagnosis of human prostate cancer and its preparation and use
  • Long-chain non-coding RNA for early diagnosis of human prostate cancer and its preparation and use

Examples

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Embodiment 1

[0082] Example 1 RACE method to obtain precise full-length cDNA sequence of lncRNA MYU

[0083] This embodiment provides a method for obtaining the precise full-length cDNA sequence of lncRNA MYU, which is the long-chain non-coding RNA of the present invention, which specifically includes the following steps:

[0084] 1. Use the extraction reagent RNAiso Plus reagent (Takara) to extract the total RNA of the prostate cancer cell line DU145 by the Trizol extraction method, and use Thermo Fisher's PrimerScript RT-PCR kit (Takara) reverse transcription kit to use oligo dT as reverse transcription Primer, reverse transcription of total RNA into cDNA.

[0085] 2. Using the cDNA in step 1 as a template, design primers 5'-F and 3'-R according to the predicted sequence (the predicted sequence is the known sequence Assembly Gene ID in the Mitranscriptome database: G032453), and the primer 5' The nucleotide sequence of -F is shown in SEQ ID NO: 10, and the nucleotide sequence of the primer 3'-...

Embodiment 2

[0093] Analysis of the conservation, transcription activity and coding ability of the long non-coding RNA described in Example 2

[0094] In this embodiment, bioinformatics algorithms are used to analyze the conservation, transcription activity, and coding ability of the long non-coding RNA (precise full-length cDNA sequence of lncRNAMYU) obtained in Example 1. The specific analysis method is as follows:

[0095] 1. Use Roadmap to obtain the data of H3K4me3, H3k36me3 and H3K27ac in the prostate cancer cell line PC3, and use the ENCODE database (https: / / www.encodeproject.org / ) to obtain the data of H3K4me3, H3k36me3 and H3K27ac in normal tissues, GEO accession number They are GSE96019 (H3K4me3, PC3), GSE96418 (H3k36me3, PC3) and GSE96399 (H3K27ac, PC3), ENCODE Experiment IDs are ENCSR748RBT (H3K4me3, Normal prostate), ENCSR499FXI (H3k36me3, Normal prostate ID) and ENCSR 76327 (Normal prostateID) , And display the obtained data on the UCSC website (http: / / genome.ucsc.edu / ) to obtain ...

Embodiment 3

[0098] Example 3 Detection of the long-chain non-coding RNA expression in different prostate cancer cell lines

[0099] This embodiment provides a specific primer for detecting the expression level of the long non-coding RNA (lncRNA MYU) in cells and tissues; the specific primer includes the upstream primer MYU- F and the downstream primer MYU-R, the nucleotide sequence of the upstream primer MYU-F is shown in SEQ ID NO. 4, and the nucleotide sequence of the downstream primer MYU-R is shown in SEQ ID NO. 5.

[0100] This embodiment provides a kit for early diagnosis of human prostate cancer, which includes the above-mentioned specific primers.

[0101] The kit for early diagnosis of human prostate cancer includes: Premix Ex Taq (TliNaseH Plus) (2×), 10 μL; the upstream primer MYU-F, 10 μM, 0.4 μL; the downstream primer MYU-R, 10 μM, 0.4 μL; ROX Reference Dye II, 0.4 μL; dH 2 O, 6.8 μL.

[0102] This embodiment provides a qRT-PCR reaction system for early diagnosis of human prostate ...

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Abstract

The invention discloses long-chain non-coding RNA for diagnosing a human prostate cancer in the early stage and preparation and application. The cDNA sequence of the long-chain non-coding RNA is shownas SEQ ID NO.1, an accurate lncRNA MYU full-length gene sequence is cloned for the first time, the foundation is laid for researching the function of the sequence in the prostate cancer occurrence and development process, high expression of the long-chain non-coding RNA in the prostate cancer is achieved through analysis, expression in normal tissue is low, and the expression of the long-chain non-coding RNA is significantly related to deterioration of the prostate cancer. Therefore, the invention further discloses the application of the long-chain non-coding RNA in preparing products for diagnosing or treating the prostate cancer, for example, a short hairpin segment of an RNAi target site of the long-chain non-coding RNA and expression of recombinant plasmid transcribing short hairpin RNA in prostate cancer cells can obviously degrade cell endogenous long-chain non-coding RNA and inhibit malignant growth and migration of the prostate cancer cells.

Description

Technical field [0001] The invention belongs to the field of molecular diagnosis of human prostate cancer, and specifically relates to a long-chain non-coding RNA used for early diagnosis of human prostate cancer, and its preparation and use. Background technique [0002] Prostate cancer (PCa) is a non-epithelial malignant tumor that occurs in the prostate, which seriously endangers men's health. Worldwide, the incidence of prostate cancer among male malignancies ranks second among all male malignancies. In developed countries such as Europe and the United States, the incidence of prostate cancer is relatively high, while the survival rate of prostate cancer patients in poor or some rural areas is low. The incidence of prostate cancer in Asia is far lower than that in European and American countries, but it has shown an upward trend in recent years, and it has grown more rapidly than in developed countries in Europe and America. At present, among the important health problems f...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/11C12N15/10C12Q1/6886A61K31/7088A61P35/00
CPCA61K31/7088A61P35/00C12N15/1096C12N15/113C12N2310/14C12N2310/531C12N2330/30C12Q1/6886C12Q2600/118C12Q2600/158C12Q2600/178C12Q2531/113
Inventor 高山汪金华
Owner SUZHOU INST OF BIOMEDICAL ENG & TECH CHINESE ACADEMY OF SCI
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