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Long-chain non-coding RNA and application thereof

A long-chain non-coding, mir-671 technology, applied in DNA/RNA fragments, recombinant DNA technology, non-central analgesics, etc., can solve problems such as limited research

Active Publication Date: 2019-06-18
NANTONG UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

LncRNA is closely related to a variety of diseases. At present, most of the research on LncRNA mainly focuses on the relationship with cancer and its role as a tumor-specific marker, while the research on LncRNA in other physiological activities or diseases is very limited.

Method used

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  • Long-chain non-coding RNA and application thereof
  • Long-chain non-coding RNA and application thereof
  • Long-chain non-coding RNA and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026]Example 1 Investigating the expression and distribution characteristics of LncRNA LOC680254

[0027] 1. RNA extraction and qRT-PCR of sciatic nerve tissue

[0028] The tissues near the sciatic nerve crush of rats on 0d, 1d, 4d, 7d, and 14d were collected and pressed Reagent (Invitrogen) instructions extract tissue RNA, after reverse transcription, use PrimeScript RT-PCR Kit (Takara) was used for qRT-PCR, and the operation was performed according to the kit instructions (GAPDH was used as an internal reference). ℃30s; Stage 3: 95℃15s, 60℃1min, 95℃15s. The primer sequence of LncRNA LOC680254 is shown in SEQ ID No: 6-7. qRT-PCR results such as figure 1 As shown in a, the results showed that the expression of LncRNA LOC680254 in the injured sciatic nerve tissue was continuously up-regulated compared with 0d.

[0029] 2. Rapid amplification of cDNA ends (RACE)

[0030] According to SMARTer TM According to RACE (Clotech) instructions, Tm>70°C, base number around 28nt...

Embodiment 2

[0033] Example 2 Investigating the effects of interfering with the expression of LncRNA LOC680254 on the proliferation, apoptosis and cell cycle of Schwann cells

[0034] 1. Culture of primary Schwann cells

[0035] Take several sciatic nerves of newborn 1-day-old SD rats, add 1ml 3mg / ml collagenase, cut up the tissue, and digest at 37°C for 30min; centrifuge at room temperature at 1200rpm for 5min, discard the collagenase, add 1ml of trypsin, and digest at 37°C for 10min ; Add 3ml of complete medium to stop digestion, filter the sieve, centrifuge to remove the supernatant (1200rpm, 5min), then add 3ml of complete medium to wash 2 times; culture the cells in a PLL-coated culture dish (5% CO 2 , 37℃); on the second day, change to the complete medium containing Arac (10μM) to inhibit the rapid proliferation of fibroblasts; on the fourth day, change to the complete medium containing HRG (50ng / ml) and Forskolin (2μM) The base stimulates the rapid growth of Schwann cells. After t...

Embodiment 3

[0047] Example 3 Investigation of LncRNA LOC680254 affecting the expression of genes Psrc1 and Ska1 by regulating miR-30d-3p, miR-671 and miRNA-3594-3p, miRNA-3473

[0048] 1. Target gene screening

[0049] The primary cultured Schwann cells were transfected with LncRNA LOC680254-specific siRNA and Negativecontrol (NC), the total RNA was extracted, and expression profiling was performed using an Agilent expression profiling chip to screen 548 genes that were down-regulated by more than 2 times. According to the full-length sequence of LncRNA LOC680254, Tay Y et al. (Tay Y, Kats L, Salmena L, et al. Coding-independent regulation of the tumor suppressor PTEN by competing endogenous mRNAs[J].Cell,2011,147(2):344 -357.) Established bioinformatics method for predicting ceRNA, and selected 191 target genes and miRNAs that LncRNA LOC680254 may regulate by ceRNA. Among the 191 target genes, 54 genes were down-regulated by more than 3 times, and 82 genes had more than 2 miRNAs binding...

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Abstract

The invention relates to long-chain non-coding RNA and an application thereof. LncRNA LOC680254 related to cell proliferation and cell cycle control is obtained by analyzing different LncRNAs in different time point samples after sciatic nerve injury of SD rats by use of RNA-seq and can be used as miRNA molecular sponge to specifically bind miR-30d-3p, miR-671, miR-3594-3p or miR-3473 so as to resist miRNA functions and promote expression of Psrc1 and Ska1, and cell proliferation is promoted.

Description

technical field [0001] The invention belongs to the professional field of biomedicine, and relates to a long-chain non-coding RNA and its application. technical background [0002] Long non-coding RNA (LncRNA) is a type of RNA with a length greater than 200nt and no coding function. LncRNA expression is spatiotemporally specific and can regulate gene expression at various levels such as chromatin remodeling, transcriptional regulation, and post-transcriptional processing. LncRNA is closely related to a variety of diseases. At present, most of the research on LncRNA mainly focuses on its relationship with cancer and its role as a tumor-specific marker, while the research on LncRNA in other physiological activities or diseases is very limited. [0003] In multicellular organisms, there is a very strict regulatory system for various cell cycles, enabling various types of cells to proliferate or remain in a quiescent state according to the needs of the body. In addition to bei...

Claims

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Application Information

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IPC IPC(8): C12N15/113A61K45/00A61P35/00A61P1/16A61P11/00A61P13/12A61P13/08A61P19/02A61P29/00A61P17/06A61P9/10A61P25/00A61P3/10A61P25/28A61P7/06A61P25/16A61P25/14A61P37/02A61P9/04A61P31/18A61P7/04A61P7/02A61P7/00
CPCA61K45/00A61P1/16A61P3/10A61P7/04A61P7/06A61P9/04A61P9/10A61P11/00A61P7/00A61P7/02A61P13/08A61P13/12A61P17/06A61P19/02A61P25/00A61P25/28A61P29/00A61P25/14A61P25/16A61P31/18A61P35/00A61P37/02C12N15/113A61K48/00
Inventor 于彬姚淳吴建成杨宇民顾晓松
Owner NANTONG UNIVERSITY
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