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Porcine endometrium epithelial cell line susceptible to PRRSV and construction method of cell line

A technology of endometrium and epithelial cells, applied in the field of transgenics, can solve the problems of inability to respond to infection and restriction of natural host cells, and achieve the effect of expanding the range and prolonging the cell cycle

Inactive Publication Date: 2019-03-19
SHANDONG AGRICULTURAL UNIVERSITY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

PRRSV has a strong tropism for host cells. The cells currently used to isolate and culture PRRSV are mainly porcine alveolar macrophages (PAM) and African green monkey kidney cells (Marc-145), but PAM is not related to the uterus and placenta cells; Marc-145 is derived from monkey rather than pig-derived cells, which will lead to the limitation of PRRSV in vitro studies that cannot reflect the infection of natural host cells

Method used

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  • Porcine endometrium epithelial cell line susceptible to PRRSV and construction method of cell line
  • Porcine endometrium epithelial cell line susceptible to PRRSV and construction method of cell line
  • Porcine endometrium epithelial cell line susceptible to PRRSV and construction method of cell line

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Embodiment 1

[0050] Example 1: Construction of porcine endometrial epithelial cell line

[0051] 1. Culture of porcine endometrial epithelial cells:

[0052] Fresh porcine uterine tissue was taken, rinsed, and the cavity surface was cut open. Rinse with PBS to remove mucus and exfoliated dead cells. The endometrium was excised and washed three times in PBS. Cut the excised endometrium into small tissue fragments (1-2mm 3 size), inoculated directly into a 6cm cell culture dish, and placed in an incubator for 15 minutes without adding culture medium. After taking it out, add a small amount of culture medium and continue to cultivate in the incubator. After four hours, add fluid until the tissue block is submerged, and add penicillin and streptomycin (penicillin 100 U / ml, streptomycin 100 μg / ml) at the same time. The medium was changed every other day, and after the porcine endometrial epithelial cells converged around the tissue block, the culture was continued for four days before use....

Embodiment 2

[0065] Example 2: Verification of the characteristics of primary porcine endometrial epithelial cells and constructed cell lines

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Abstract

The invention discloses a porcine endometrium epithelial cell line susceptible to PRRSV and a construction method of the cell line. The porcine endometrium epithelial cell line is obtained by taking porcine endometrium epithelial cells as host cells, transfecting lentivirus expression plasmids with SV40T antigen genes, and performing Puromycin drug screening and cell expansion culture. The porcineendometrial epithelial cell line has the function of infinite passage, can maintain the characteristics of the porcine endometrial epithelial cells, can maintain normal karyotype in the passage process, and does not have changes in chromosome structure and number; and moreover, the porcine endometrial epithelial cell line is sensitive to PRRSV virus, and can be used for the passage of PRRSV and related research.

Description

technical field [0001] The invention relates to the field of transgenic technology, in particular to a pig endometrium epithelial cell line susceptible to PRRSV and a construction method thereof. Background technique [0002] Porcine reproductive and respiratory syndrome, also known as porcine blue ear disease, causes huge losses in the production process of pigs. The disease is caused by porcine reproductive and respiratory syndrome virus (PRRSV). The disease only infects pigs, other animals will not be infected with the virus, and pigs of various ages and breeds can be infected with the virus, among which pregnant sows and piglets within one month are the most susceptible. Clinically, the characteristic symptoms are abortion of pregnant sows and premature delivery of fetuses. Previous studies have shown that fetuses infected with PRRSV will be aborted due to the apoptosis of placental cells. The endometrium and fetal placenta play an important role in the establishment ...

Claims

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Application Information

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IPC IPC(8): C12N5/10C12N15/867C12N7/00C12R1/93
CPCC12N5/0682C12N7/00C12N15/86C12N2510/00C12N2740/15043C12N2770/10051C12N2800/107
Inventor 苏峰张康姜运良葛利江刘海昌胡栋王栋刘营张红焱
Owner SHANDONG AGRICULTURAL UNIVERSITY
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