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Long-chain non-coding RNA marker for diagnosing lung adenocarcinoma

A long-chain non-coding, lung adenocarcinoma technology, applied in the field of biomedicine, can solve the problem of unknown function of lncRNA, and achieve the effect of providing survival rate

Active Publication Date: 2017-08-18
FOURTH HOSPITAL OF HEBEI MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, more than 40,000 human lncRNAs have been cloned, but the functions of most lncRNAs are unknown

Method used

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  • Long-chain non-coding RNA marker for diagnosing lung adenocarcinoma
  • Long-chain non-coding RNA marker for diagnosing lung adenocarcinoma
  • Long-chain non-coding RNA marker for diagnosing lung adenocarcinoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1 Screening of differentially expressed non-long-chain coding RNA

[0050] 1. Materials:

[0051] The surgically resected cancer tissues and paracancerous tissues of 8 patients with primary lung adenocarcinoma were used as experimental samples. All cancer tissues were pathologically confirmed as lung adenocarcinoma. All patients with primary lung adenocarcinoma had not received radiotherapy and chemotherapy before operation, and the clinical data of all cases were complete.

[0052] 2. Obtaining tissue RNA

[0053] Total RNA was extracted from tissue samples, and the concentration and purity of the extracted RNA were detected by Nanodrop2000, the integrity of RNA was detected by agarose gel electrophoresis, and the RIN value was determined by Agilent2100. The total amount of RNA required for a single library construction is 5ug, the concentration is ≥200ng / μL, and the OD260 / 280 is between 1.8 and 2.2.

[0054] 3. Remove rRNA

[0055] A part (>24%) of the lo...

Embodiment 2

[0079] Example 2 Large sample verification screened out differentially expressed genes

[0080] Based on the results of previous high-throughput sequencing and according to the size of P value, we selected LOC102724660 for verification.

[0081] 1. Sample collection

[0082] According to the method of Example 1, 45 cases of lung adenocarcinoma tissues and corresponding paracancerous tissues were collected.

[0083] 2. Validation at the mRNA level

[0084] Reagents: Reverse transcription kit (DDR037A) was purchased from Treasure Bioengineering (Dalian) Co., Ltd. SYBR Premix ExTaq for real-time quantitative PCR (polymerase chain reaction) TM (Tli RNaseHPlus) kit was produced by Japan Takara Company.

[0085] 2.1 Extract tissue RNA

[0086] Step is with embodiment 1.

[0087] 2.2 Primer design

[0088] According to the sequences of the five transcripts of LOC102724660, primers were designed by NCBI's primer design tool (PrimerBLAST). The designed primers were suitable for ...

Embodiment 3

[0095] Example 3 Overexpression of LOC102724660

[0096] 1. Plasmid construction

[0097] The cDNA of LOC102724660 was obtained by the method of Example 2, and the amplified sequence was the cDNA sequence of LOC102724660 shown in SEQ ID NO.1. The above cDNA sequence was inserted into the eukaryotic cell expression vector pcDNA3.1, and the obtained recombinant vector pcDNA3.1-LOC102724660 was connected for subsequent experiments.

[0098] 2. Culture and transfection of lung adenocarcinoma cells

[0099] 2.1 Cell culture

[0100] The lung adenocarcinoma cell line A549 was cultured in RPMI1640 medium and 10% fetal bovine serum.

[0101] 2.2 Cell transfection

[0102] (1) The day before transfection, 0.5-2*10 5 Tumor cells were suspended in 500 μl of antibiotic-free medium and seeded into 24-well culture plates.

[0103] (2) On the day of transfection, the cell density should reach 80%-90%. Prepare the following complex A: Dilute 1 μg plasmid DNA in serum-free medium and mix...

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Abstract

The invention provides long-chain non-coding RNA and application thereof. Studies suggest that expression of LOC102724660 in lung adenocarcinoma tissue is obviously different from expression in para-carcinoma tissue. On this basis, it is concluded that LOC102724660 can serve as a molecular marker for diagnosing lung adenocarcinoma. Based on the fact that LOC102724660 is expressed at a low level in lung adenocarcinoma tissue and is over-expressed in lung adenocarcinoma cells, the function of LOC102724660 in lung adenocarcinoma is researched, and the result show that overexpression of LOC102724660 can inhibit the proliferation of lung adenocarcinoma cells remarkably. The key role of LOC102724660 in the lung adenocarcinoma pathogenesis is disclosed, and a novel molecular marker and drug target are provided for clinical diagnosis, treatment and prognosis monitoring of lung adenocarcinoma.

Description

technical field [0001] The invention belongs to the field of biomedicine and relates to a long-chain non-coding RNA marker for diagnosing lung adenocarcinoma. Background technique [0002] There are about 20,000 to 30,000 genes that encode proteins in the human body, accounting for only 2% of the human genome, and the remaining 98% of genomic DNA that does not encode proteins was originally considered to be nonfunctional and garbage in organisms, often called "Junk DNA". [0003] However, current research shows that most of these junk DNA can be transcribed to produce non-coding RNA (non-coding RNA, ncRNA). According to the size of mature transcripts, ncRNA can be divided into small molecule ncRNA (such as siRNA, miRNA, piRNA, etc.), medium-length ncRNA (70-200nt) and long ncRNA (long ncRNA, LncRNA, >200nt). [0004] At present, most of the research in the field of ncRNA is small molecule ncRNA, while the research on lncRNA is still in its infancy. Due to the presence ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68A61K45/00A61P35/00
CPCA61K45/00C12Q1/6886C12Q2600/118C12Q2600/136C12Q2600/158C12Q2600/178
Inventor 田子强李振华温士旺
Owner FOURTH HOSPITAL OF HEBEI MEDICAL UNIV
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