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Fibroblast cell line of Hubei white pig fetus

A technology of fibroblasts and fetuses, applied in the field of cell biology, can solve the problems of animal husbandry loss, shrinkage, and affecting the survival of breeds

Active Publication Date: 2014-01-15
INST OF ANIMAL SCI & VETERINARY HUBEI ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, in many years of production, due to factors such as overuse of Hubei white pigs and ignorance of the original breed, the excellent genetic resources of Hubei white pigs have not been effectively promoted, and even have a tendency to shrink in recent years, which directly affects the development of this breed. Survival will cause immeasurable losses to my country's animal husbandry

Method used

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  • Fibroblast cell line of Hubei white pig fetus
  • Fibroblast cell line of Hubei white pig fetus
  • Fibroblast cell line of Hubei white pig fetus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Example 1 Isolation and Primary Culture of Hubei White Pig Fetal Fibroblasts.

[0051] (1) Material

[0052]

[0053] (2) Method

[0054] The fetuses of Hubei white pigs at 35 days of pregnancy were taken out during the operation, washed 6-8 times with normal saline, and the residual blood and body fluids were washed away as much as possible to prevent contamination by bacteria and viruses.

[0055] Use tweezers and scissors in the ultra-clean workbench to cut the body of the fetus into pieces, and discard the internal organs, head, eyes and other tissues and organs. Only the tissues of the back and buttocks are kept and cut into fine pieces.

[0056] Transfer the tissue pieces to a fresh centrifuge tube, add an appropriate amount of saline, and grind them into pieces as much as possible with tweezers while washing. Repeat 3 times.

[0057] Add trypsin solution and let it stand overnight at 4°C to allow trypsin to digest connective tissue, etc.

[0058] The next ...

Embodiment 2

[0062] Example 2 Subculture, cryopreservation, recovery and survival rate determination of Hubei white pig fetal fibroblasts.

[0063] (1) Material

[0064]

[0065] (2) Method

[0066] After the primary cells covered the integrated culture plate, they were digested with trypsin solution according to 10 4 The density of cells / ml was seeded on the corresponding cell culture plate and returned to the incubator for culture. This is the inheritance.

[0067] After the cells cover the bottom of the culture plate, digest it with trypsin solution to form a single-cell suspension, transfer it to a freshly sterilized 1.5ml centrifuge tube, collect the cells at the bottom of the tube in a desktop centrifuge at 6000rpm for 2min, and remove the supernatant completely , add cell freezing solution. The formula of the cryopreservation solution is DMEM: fetal bovine serum: DMSO=6:3:1. Still mix the cells, wrap them in cotton, freeze them overnight in a -80°C ultra-low temperature refr...

Embodiment 3

[0070] The doubling time measurement of embodiment 3 Hubei white pig fetal fibroblasts

[0071] (1) Material

[0072]

[0073] (2) Method

[0074] The vigorously growing Hubei white pig fetal fibroblasts were prepared according to 10 4 The density of cells / ml was inoculated on a 24-well plate, and a total of 3 × 6 = 21 wells were inoculated, and then returned to the incubator for culture.

[0075] From the next day (ie, after 24 hours), the cells in the 3 wells were digested with trypsin solution every other day to form a single cell suspension, and the cell density was counted with a red blood cell counting board. Each count corresponds to 3 biological replicates.

[0076] The data of 6 experiments were input into the Prism5 software to draw the growth curve of the cells.

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Abstract

The invention belongs to the field of cell biology. A morphologically uniform and vigorously-growing fibroblast cell line of Hubei white pig fetus is prepared through the following steps: taking a Hubei white pig fetus which has been in pregnancy for 35 days as the raw material, then subjecting the fetus to treatments of mechanical shearing and pancreatin digestion, and finally subjecting the digestion product to processes of primary culture, subculture, liquid nitrogen freezing storage, and cell thawing so as to obtain the finished product; the fibroblast cell line is preserved in the China Center for Type Culture Collection (CCTCC), and the preservation number is CCTCC No. C201331. The obtained fibroblast cell line of Hubei white pig fetus has a high purity, a uniform shape, and no compounding of epithelial cells, liver cells, myocardial cells, and the like. The cell line provided by the invention can provide high quality cell materials for gene engineering, cell engineering, embryo engineering, immunology and molecular biology, can be used as donor cells for porcine somatic cell nuclear transplantation, is an important genome resource bank, and is a species preservation method for local excellent variety.

Description

technical field [0001] The invention relates to a Hubei white pig fetal fibroblast cell line and related biological characteristics, belonging to the field of cell biology. Background technique [0002] Biological genetic resources are the foundation and foundation of the existence and development of human society, and are strategic resources for achieving sustainable social development and maintaining national food, ecological security and social stability. In recent years, due to the continuous deterioration of the earth's ecological environment, biological genetic resources are facing a serious crisis, and some are even on the verge of exhaustion and extinction. The protection, development and utilization of biological genetic resources have attracted great attention from governments all over the world. Saving endangered species, preserving genetic resources, and strengthening development and utilization have become top priorities. [0003] Our country has a vast territ...

Claims

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Application Information

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IPC IPC(8): C12N5/073C12N15/873C12R1/91
Inventor 毕延震刘西梅肖红卫郑新民华文君张立苹
Owner INST OF ANIMAL SCI & VETERINARY HUBEI ACADEMY OF AGRI SCI
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