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Method for coustructing shRNA expression carrier for inhibiting N-acetylglucosamine transferase V and its use

A technology of acetylglucosamine and its construction method, which is applied in the field of specific small molecule hairpin interfering RNA, which can solve the problems of large side effects, high cost, and difficult operation, and achieve the effects of small side effects, convenient application, and easy transfection

Inactive Publication Date: 2009-05-13
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, some researchers have used swainsonine to inhibit the expression of Mgat5 in mice, and there are similar changes
However, these two approaches have disadvantages such as difficult operation, high cost and large side effects.

Method used

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  • Method for coustructing shRNA expression carrier for inhibiting N-acetylglucosamine transferase V and its use
  • Method for coustructing shRNA expression carrier for inhibiting N-acetylglucosamine transferase V and its use
  • Method for coustructing shRNA expression carrier for inhibiting N-acetylglucosamine transferase V and its use

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Experimental program
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Embodiment Construction

[0054] 1. Design primers and construct vectors ( figure 1 ).

[0055] A. According to the principle of siRNA target sequence selection, according to the mouse MGAT5 gene sequence (AF474155) provided by Genebank, select the 836th-856th nucleotide sequence TCGGGCTTCAAGATTGCGG from the coding region as the target sequence, design and synthesize primer design, synthesis Primers (synthesized by Shanghai Saibaisheng Co., Ltd.), the primers are:

[0056] siRNA1-Oligo1: 5'-TCGGGCTTCAAGATTGCGGTTCA-3'

[0057] siRNA1-Oligo2: 5'-AGCTTGAACCGCAATCTTGAAGCCCGAGGCC-3'

[0058] siRNA1-Oligo3: 5'-AGCTTCCGCAATCTTGAAGCCCGATTTTTT-3'

[0059] siRNA1-Oligo4: 5'-AATTAAAAAATCGGGCTTCAAGATTGCGGA-3'

[0060] siRNA2-Oligo1: 5'-AAGATTGAGCCGTATATGCCATTCA-3'

[0061] siRNA2-Oligo2: 5'-AGCTTGAATGGCATATACGGCTCAATCTTGGCC-3'

[0062] siRNA2-Oligo3: 5'-AGCTTTGGCATATACGGCTCAATCTTTTTT-3'

[0063] siRNA2-Oligo4: 5'-AATTAAAAAAGATTGAGCCGTATATGCCAA-3'

[0064]B. Digest the vector psilencer1.0-U6 (Ambion, USA) w...

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Abstract

The invention discloses a construction method for shRNA eukaryon expression vector that could restrain N-acet glucose amine transferase V and the application. The process includes the following steps: selecting the 836th to 856th nucleotide sequence TCGGGCTTCAAGATTGCGG and designing as target sequence and compounding to primer according to the gene sequence of MGAT5 supplied by Genebank, taking primer annealing and connecting to carrier psilencer1.0-U6 to gain eukaryon expression shRNA1-Mgat5 that would be transfected to breast cancer cell MA782 to restrain the growth in mousebody. The application of the particle has wide prospects in medicinal to cure or protect breast cancer.

Description

technical field [0001] The present invention belongs to the field of molecular biology. More specifically, it relates to a method for constructing an shRNA eukaryotic expression vector that inhibits N-acetylglucosamine transferase V, and also relates to a method that can specifically target and inhibit mouse N-acetylglucosamine transferase V (Mouse Golgi β1, 6N-acetylglucosaminyltransferase V, also known as Mgat5) gene expression specific small hairpin interfering RNA (ShRNA, Small hairpin RNA), in vivo inhibition of breast cancer cells purposes. Background technique [0002] Breast cancer is a malignant tumor that seriously endangers women's health. It has become the first cause of death of women in western developed countries, and it is also one of the nine common malignant tumors in my country. 9th among all cancers. With the large-scale prospective clinical research on various stages of breast cancer and the continuous understanding of the biology of breast cancer, it i...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/79C07H21/04C12N1/19A61K48/00A61P35/00
Inventor 章晓联李冬青
Owner WUHAN UNIV
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