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Building method of donor pig suitable for kidney xenotransplantation
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A technology for xenotransplantation, construction method, applied in the field of construction suitable for kidney xenotransplantation donor pigs
Inactive Publication Date: 2019-10-25
YUNNAN AGRICULTURAL UNIVERSITY
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However, using GGTA1 knockout (GTKO) pigs for xenotransplantation of pigs to non-human primates such as hearts and kidneys, although GTKO overcomes hyperacute immune rejection to a certain extent, the recipients eventually suffer from acute vascular rejection (AVR). ) caused by thrombotic microangiopathy
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Embodiment 1
[0022] A construction method suitable for kidney xenotransplantation donor pigs, the specific operation process is carried out according to the following steps:
[0024] On the basis of porcine fibroblastcell lines, three antigen genes that cause immune rejection, GGTA1, CMAH, and B4GalNT2, were knocked out using CRISPR / Cas9 gene editing technology;
[0025] 2) Site-specific insertion of three humanized modification genes of hCD55, hCD59, and hCD46
[0026] On the basis of the above-mentioned GTKO / CMAHKO / B4GalNT2KO three-gene modification, three humanized modification genes of hCD55, hCD59, and hCD46 were inserted at specific sites to construct porcine fibroblasts with GTKO / CMAHKO / B4GalNT2KO / hCD55 / hCD59 / hCD46 six-gene modification Tie;
[0031] A construction method suitable for kidney xenotransplantation donor pigs, the specific operation process is carried out according to the following steps:
[0032] 1) Knockout of the three major antigen genes that cause immune rejection
[0033] On the basis of porcine fibroblastcell lines, three antigen genes that cause immune rejection, GGTA1, CMAH, and B4GalNT2, were knocked out using CRISPR / Cas9 gene editing technology;
[0034] 2) Site-specific insertion of three humanized modification genes of hCD55, hCD59, and hCD46
[0035] On the basis of the above-mentioned GTKO / CMAHKO / B4GalNT2KO three-gene modification, three humanized modification genes of hCD55, hCD59, and hCD46 were inserted at specific sites to construct porcine fibroblasts with GTKO / CMAHKO / B4GalNT2KO / hCD55 / hCD59 / hCD46 six-gene modification Tie;
[0037] Using somatic cell nuclear transfer technology, the screened GTKO / CMAHKO / B4GalNT2KO / hCD5...
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Abstract
The invention relates to a building method of a donor pig suitable for kidneyxenotransplantation and belongs to the technical field of animal biology. The building method includes: on the basis of porcine fetal fibroblast, utilizing CRISPR / Cas9 gene editing technology to knock out three genes GGTA1, CMAH and B4GalNT2 at the same time, and inserting three humanized modifying genes hCD55, hCD59 andhCD46 in a fixed point manner; combining somatic cellnuclear transfer technology to build an xenotransplantation donor base pig modified by six genes GTKO / CMAHKO / B4GalNT2KO / hCD55 / hCD59 / hCD46; on thebasis of the base pig, inserting three modifying genes hCD39, hTBM and REN in a fixed point manner to build a kidneyxenotransplantation donor pig modified by multiple genes; performing functional verifying on the donor pig to obtain the donor pig suitable for kidney xenotransplantation. A scientific theoretic basis is provided for developing the donor pig suitable for xenotransplantation, and hope is provided to transplantation therapy of patients with kidney failure.
Description
technical field [0001] The invention belongs to the field of animal biotechnology, and in particular relates to a method for constructing kidney xenotransplantation donor pigs. Background technique [0002] Organ transplantation is the only way to treat patients with end-stage organ failure, and xenogeneic kidney transplantation is the most effective way to solve the severe shortage of human kidney donors. In recent years, with the substantial increase of patients with chronic diseases such as cardiovascular diseases in my country, the number of patients developing end-stage heart failure is increasing sharply, and the pressure on organ demand will gradually increase. Since 2015, my country has completely stopped using the organs of executed prisoners as a source of transplant donors, and can only use organs donated voluntarily after death as the final source of organ transplants in my country. This will inevitably lead to the growing shortage of organ donors such as kidneys...
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