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Macrophage specific targeting vector and recombinant cell constructed based on Ipr1 gene

A technology for targeting vectors and macrophages, applied to cells modified by introducing foreign genetic material, genetic engineering, plant genetic improvement, etc.

Active Publication Date: 2012-06-27
NORTHWEST A & F UNIV +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the current treatment of tuberculosis can only be treated with antibiotics, such as: streptomycin, kanamycin, etc.

Method used

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  • Macrophage specific targeting vector and recombinant cell constructed based on Ipr1 gene
  • Macrophage specific targeting vector and recombinant cell constructed based on Ipr1 gene
  • Macrophage specific targeting vector and recombinant cell constructed based on Ipr1 gene

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Embodiment Construction

[0042] The present invention first amplifies the MSR1 promoter and the homologous long and short arms from the bovine blood genome, and connects the Ipr1 gene and the above fragments to the universal targeting vector pA2T to construct the macrophage-specific targeting vector pLMIS, and then use The linearized pLMIS was introduced into neonatal bovine ear fibroblasts by electrotransfection, and positive cell clones were obtained by G418 and GCV positive and negative screening. PCR and Southern blot identification confirmed that the target gene Ipr1 was integrated into the neonatal bovine ear fibroblasts. The test results of the generated cloned cattle indicate that the obtained cattle are transgenic cloned cattle with Ipr1 site-specific insertion, and can resist M. bovis infection.

[0043] In the following, the present invention will be further described in detail with reference to the drawings and experiments, which is to explain but not limit the present invention.

[0044] 1. Co...

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Abstract

The invention discloses a macrophage specific targeting vector and a recombinant cell constructed based on an Ipr1 gene. According to the invention, an MSR1 promoter is connected to a 5' terminal of the Ipr1 gene, such that the Ipr1 gene is specifically expressed in bovine macrophage with high efficiency; a recombinant gene LA-MSR1-Ipr1-SA is further constructed, such that a target gene is subject to site-directed integration, and is integrated between bovine 28-chromosome SP-A and MAT1A gene. An Ipr1 macrophage specific targeting vector pLMIS is constructed, and is transfected into newborn bovine ear fibroblast cells; through G418 and GCV positive and negative screening, transgenic newborn bovine ear fibroblast cell lines are constructed; the transgenic newborn bovine ear fibroblast celllines are adopted as nuclear donors, and are transplanted into bovine enucleated oocytes, such that transgenic cloned embryos are obtained; when the embryos are transplanted, five cattle with site-directed inserted Ipr1 gene is produced, and four of them survive. The transgenic cattle is resistant to M.bovis.

Description

Technical field [0001] The invention belongs to the technical field of transgenic cloned animals, and relates to a macrophage heterogeneous expression vector and recombinant cells constructed based on the Ipr1 gene. Background technique [0002] Bovine tuberculosis is a zoonotic disease caused by M.bovis, and it is one of the most catastrophic diseases of cattle in developing countries (Berrada and Barjas-Rojas, 1995). Bovine Mycobacterium tuberculosis mainly affects cattle. Bovine tuberculosis is a zoonotic chronic infectious disease caused by Mycobacterium bovis. Its pathological changes are characterized by the gradual weight loss of sick cattle and the formation of nodular granulation in tissues and organs. Swollen and caseous necrosis. Bovine tuberculosis is easily transmitted and has serious harm. Therefore, the World Animal Health Organization lists it as a Class B infectious disease, and my country lists it as a Class II animal infection. Its spread affects the sustaina...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12N15/85C12N5/10C12N15/877
Inventor 何小宁张涌权富生王勇胜
Owner NORTHWEST A & F UNIV
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