siRNA (small interfering Ribonucleic acid) for specifically inhibiting XOR (oxidoreductase) gene expression and application of siRNA

A gene expression, species-specific technology, applied in the field of siRNA

Active Publication Date: 2014-12-24
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] Whether XOR plays a regulatory role in NAFLD has not been reported in the literature

Method used

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  • siRNA (small interfering Ribonucleic acid) for specifically inhibiting XOR (oxidoreductase) gene expression and application of siRNA
  • siRNA (small interfering Ribonucleic acid) for specifically inhibiting XOR (oxidoreductase) gene expression and application of siRNA
  • siRNA (small interfering Ribonucleic acid) for specifically inhibiting XOR (oxidoreductase) gene expression and application of siRNA

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Embodiment 1, siRNA design and synthesis

[0032] Obtain the full-length sequence of human XOR mRNA (NM_000379.3) from the National Center for Biotechnology Information (NCBI) database, according to the principle of RNAi, combined with design software, and on the basis of experiments, design and synthesize and screen for 4 transcripts An effective 21nt siRNA against human XOR gene was produced. A BLAST comparison check was performed to ensure that there was no homology to other genes.

[0033] The XOR siRNA sequence is:

[0034] 5'-CGGAAGAGUGAGGUUGACAAGUUCA-3' (sense strand)

[0035] 5'-UGAACUUGUCAACCUCACUCUUCCG-3' (antisense strand)

[0036] Chemically synthesized by Shanghai Yingwei Jieji Trading Co., Ltd. (factory address: 1711, Nanyin Building, No. 2 Dongsanhuan North Road, Shanghai, Zip Code: 200051): using NTP as raw material, using ABI3900 nucleic acid synthesizer to chemically synthesize single-stranded RNA , and finally each single-stranded RNA is anneale...

Embodiment 2

[0037] Example 2, the effect of XOR siRNA on the expression of XOR gene in L02 cells in vitro

[0038] 1. Group transfection

[0039] Spread L02 cells evenly in a 6-well plate and divide them into 4 groups:

[0040] High-fat experiment group: DMEM / F12 medium containing 10% (volume percentage) inactivated neonatal bovine serum, 666 μmol / L sodium oleate and 333 μmol / L sodium palmitate; transfection 200 pmol Example 1 XOR siRNA.

[0041] High-fat control group: DMEM / F12 medium containing 10% (volume percentage) inactivated neonatal bovine serum, 666 μmol / L sodium oleate and 333 μmol / L sodium palmitate; transfection 200 pmol Example 1 negative control siRNA.

[0042] Normal experimental group: DMEM / F12 medium containing 10% (volume percent) inactivated neonatal bovine serum, 100 U / ml penicillin and 100 mg / ml streptomycin; transfected with 200 pmol of the XOR siRNA of Example 1.

[0043] Normal control group: DMEM / F12 medium containing 10% (volume percentage) inactivated neon...

Embodiment 3

[0048] Example 3, the effect of XOR siRNA on L02 cell lipid change in vitro

[0049] 1. Group transfection

[0050] The L02 cells were spread evenly in a 6-well plate and divided into 4 groups. Concrete grouping is with the step one of embodiment 2.

[0051] 2. The effect of XOR siRNA on lipid change of L02 cells in vitro

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Abstract

The invention discloses siRNA small interfering Ribonucleic acid) for specifically inhibiting XOR (oxidoreductase) gene expression and application of siRNA. The siRNA for specifically inhibiting XOR gene expression is a double-chain RNA which is composed of nucleotide sequences as shown in SEQ ID No.1 and SEQ ID NO.2; the siRNA is modified to obtain modified RNA. According to in-vitro test results, the RNA provided by the invention can be used for specifically and efficiently inhibiting the XOR gene expression, lowering lipid droplets and inhibiting the deposition of triglyceride. The siRNA can be applied to pathogenesis research of non-alcoholic fatty liver diseases and potential treatment of the non-alcoholic fatty liver disease. The siRNA has important value.

Description

technical field [0001] The invention relates to a siRNA specifically inhibiting the expression of XOR gene and its application. Background technique [0002] RNA interference (RNA interference, RNAi) is an evolutionarily conserved defense mechanism against the invasion of transgenes or foreign viruses. The process in which double-stranded RNA (dsRNA) specifically degrades the mRNA in the cell, resulting in the effective closure of specific genes, is a sequence-specific post-transcriptional gene silencing (PTGS). RNAi technology has been successfully applied to the study of gene functions in various organisms. Construct siRNA for the target gene, use RNAi technology to shut down the expression of the gene, and analyze the function of the gene according to the change of the phenotype. Gene knockout technology takes a long time to permanently shut down the expression of a certain gene, while RNAi technology can controllably shut down 10 genes within 1 week or even 2 days, so ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113A61K48/00A61P1/16C12Q1/68C12Q1/02
Inventor 虞朝辉万星勇徐承富余伟来厉有名
Owner ZHEJIANG UNIV
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