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siRNA interfering Mroh7 gene expression and application, interfering method and drug thereof

A gene expression and drug technology, applied in the field of molecular biotechnology and genetic engineering, can solve the problem of unclear function of Mroh7, and achieve the effect of improving cell viability

Active Publication Date: 2019-06-14
HENAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in rats, the function of Mroh7 is unclear

Method used

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  • siRNA interfering Mroh7 gene expression and application, interfering method and drug thereof
  • siRNA interfering Mroh7 gene expression and application, interfering method and drug thereof
  • siRNA interfering Mroh7 gene expression and application, interfering method and drug thereof

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Experimental program
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Effect test

Embodiment 1

[0045] This embodiment proposes siRNA that interferes with Mroh7 gene expression, that is, at least one of siRNA1, siRNA2 and siRNA3; the base sequence of siRNA1 is shown in SEQ ID NO.1, and the base sequence of siRNA2 is shown in SEQ ID NO.2. The base sequence of siRNA3 is shown in SEQ ID NO.3.

[0046] Search the AA sequence according to the Mroh7 mRNA sequence of the rat and record the 19 nucleotides adjacent to the 3' end of each AA base fragment, and screen the siRNA sequences with a GC content between 30% and 55%. Then make further screening according to the basic design principles of siRNA, and at the same time search the homology of the screened siRNA sequence in the genome database of GenBank with BLAST, select the sequence with more than 3 base mismatches with the non-homologous gene, and The possibility of non-specific inhibition was excluded, and three Mroh7 siRNA sequences were finally determined on the basis of meeting the above conditions.

[0047] In this embo...

Embodiment 2

[0053] The present embodiment proposes a method for interfering with Mroh7 gene expression, the steps are as follows:

[0054] 1.1 Dilute the siRNA that interferes with the normal expression of the Mroh7 gene with a serum-free medium to prepare an siRNA solution with a concentration of 10 μg / μL.

[0055] 1.2 Dilute the liposomes with serum-free medium to prepare a liposome solution with a concentration of 20 μg / μL;

[0056] 1.3 Mix the siRNA solution and the liposome solution to prepare a liposome-siRNA complex solution, and let it stand at room temperature for 20 minutes;

[0057] 1.4 In a 6-well culture plate, culture BRL-3A cells with 2 mL / well of complete medium, and the concentration of cells in each culture well is 3×10 5 a / mL;

[0058] 1.5 at 37°C, 5% CO 2 Carry out cell culture under the condition of saturated humidity until the cultured cells cover 40%-60% of the bottom area of ​​the plate;

[0059] 1.6 Discard the medium in the culture well, and wash the cells 1-...

Embodiment 3

[0064] The present embodiment proposes a method for interfering with Mroh7 gene expression, the steps are as follows:

[0065] 1.1 Dilute the siRNA that interferes with the normal expression of the Mroh7 gene with a serum-free medium to prepare an siRNA solution with a concentration of 100 μg / μL.

[0066] 1.2 Dilute the liposomes with serum-free medium to prepare a liposome solution with a concentration of 500 μg / μL;

[0067] 1.3 Mix the siRNA solution and the liposome solution for 10 minutes to prepare a liposome and siRNA complex solution;

[0068] 1.4 Cultivate BRL-3A cells in the culture wells of the 6-well cell culture plate with 2 mL / well of complete medium, and the concentration of cells in each culture well is 6×10 5 a / mL;

[0069] 1.5 at 37°C, 5% CO 2 Carry out cell culture under saturated humidity conditions until the cultured cells cover 40%-60% of the plate area;

[0070] 1.6 Discard the medium in the culture well, and wash the cells 1-2 times with serum-free c...

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Abstract

The invention proposes siRNA interfering Mroh7 gene expression and application, an interfering method and a drug thereof and belongs to the field of molecular biotechnology and genetic engineering. The siRNA is transfected in cells to regulate the Mroh7 gene expression, further affect the cell viability and regulate the normal expression of cell proliferation and apoptosis-related genes. Therefore, the siRNA interfering the Mroh7 gene expression is applied to the preparation of a drug for improving cell viability and regulating cell division and proliferation and apoptosis, and has high practical application value.

Description

technical field [0001] This application relates to the fields of molecular biotechnology and genetic engineering, that is, siRNA that interferes with Mroh7 gene expression and its application, interference method and medicine. Background technique [0002] The liver is the hub and leading organ of various substance metabolism, energy conversion and supply in the human body, and is the "distributing center" of various important information regulation molecules in the body. The liver is also an organ with a strong regenerative ability in the human body. The research on the genes related to its repair and regeneration is of great significance in the proliferation of liver cells, the treatment of diseases and the research and development of drugs. The use of RNAi technology to specifically introduce siRNA fragments into mammalian cells can reduce the normal expression of target genes, thereby reducing the expression of target proteins, so as to achieve efficient and specific gen...

Claims

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Application Information

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IPC IPC(8): C12N15/113A61K31/7115A61P43/00
Inventor 王棋文白俊杰张春艳王改平叶丙雨
Owner HENAN NORMAL UNIV
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