Method for establishing pancreatic cancer model in wild-type adult mouse through lentiviruses

A technology of lentiviral vector and pancreatic cancer, applied in virus/bacteriophage, biochemical equipment and methods, virus, etc., to achieve the effect of simple and quick operation, low requirements for experimental conditions, and overcoming time-consuming effects

Active Publication Date: 2018-03-13
成都豆麦科技有限公司
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Problems solved by technology

[0005] The purpose of the present invention is to address the deficiencies of the existing mouse models of pancreatic cancer, and to provide a method for establishing an animal model of pancreatic cancer in wild-type adult mice with a short induction period, a high and stable induction rate, and simple and convenient operation

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  • Method for establishing pancreatic cancer model in wild-type adult mouse through lentiviruses
  • Method for establishing pancreatic cancer model in wild-type adult mouse through lentiviruses
  • Method for establishing pancreatic cancer model in wild-type adult mouse through lentiviruses

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Embodiment Construction

[0034] Below in conjunction with specific embodiment, further illustrate the present invention.

[0035] 1. Construction of lentiviral vector

[0036] According to the analysis of the TCGA database, oncogenes and tumor suppressor genes with a high mutation rate in human pancreatic cancer were selected, and pTomo empty vector was used as the starting plasmid, and a lentiviral vector expressing oncogenes and silencing tumor suppressor genes was constructed through a series of conventional enzyme-cut ligation .

[0037] 2. Virus packaging, titer determination and in vitro verification

[0038] Co-transfect HEK293T cells with the constructed lentiviral plasmids and packaging plasmids pCMV8.9 and pMD2.G, replace the new medium 6 hours after transfection, collect the virus supernatant at 48h and 72h respectively, and filter at 0.45um to remove cell debris Afterwards, use 25000rpm, 4 ℃ centrifuge 2.5h to concentrate virus, PBS resuspend virus; Use QPCR method to detect virus titer,...

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Abstract

The invention discloses a method for establishing a pancreatic cancer model in a wild-type adult mouse through lentiviruses. The method comprises lentiviral vector construction, virus packaging, titerdetermination and in-vitro verification, virus injection, and pathological identification. The method comprises selecting an oncogene Kras with the highest variation rate of the human pancreatic cancer and cancer suppressor genes of Tp53, Cdkn2a and Cdkn2b, overexpressing the oncogene KrasG12D in the same lentiviral vectors, simultaneously, knocking down Tp53, Cdkn2a and Cdkn2b through shRNA, carrying out virus injection, and after 40 days, detecting significant pancreatic tumors in the adult mouse which is killed by the pancreatic cancer. The method has a short induction period and high andstable incidence, is easy to operate and fast builds the adult mouse pancreatic cancer model. The pancreatic cancer model simulates the genetic mechanism of human pancreatic cancer to the maximum extent, can be used to study the pathogenesis of human pancreatic cancer and the screening of new therapeutic drugs and therapeutic target points and provides a reference for the treatment on human pancreatic cancer.

Description

technical field [0001] The present invention relates to a method for using lentivirus to establish a pancreatic cancer model in wild-type adult mice, in particular to a method for using lentivirus to mediate the overexpression of oncogenes and the knockdown of tumor suppressor genes to induce mouse pancreatic cancer models , belongs to the technical field of animal model establishment. Background technique [0002] Pancreatic cancer is a highly malignant tumor of the digestive tract that is difficult to diagnose and treat. The 5-year survival rate is about 5%, and it is one of the malignant tumors with the worst prognosis. Although much has been done on genetic variation in pancreatic cancer, the genetic variation that induces pancreatic carcinogenesis in adults remains unknown. Considering that the main cause of human cancer (including pancreatic cancer) is the accumulation of genetic mutations in adulthood, tumor models induced in adult animals by genetic changes can bett...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/867C12N7/01A01K67/027
CPCA01K67/0278A01K2207/15A01K2217/072A01K2227/105A01K2267/0331C07K14/4703C07K14/4746C12N7/00C12N15/86C12N2740/15021C12N2740/15043
Inventor 赵旭东涂秋
Owner 成都豆麦科技有限公司
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