Construction method for anabarilius grahami cardiac cell line

A construction method and a technology for the whitefish, applied in the field of freshwater biological cell culture, can solve the problems such as the same reports that have not been invented, and achieve the effect of good growth state

Active Publication Date: 2013-11-20
KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI
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Problems solved by technology

[0003] Through document retrieval, do not see identical report with the present invention

Method used

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Embodiment Construction

[0017] The method for constructing the heart cell line of the whitefish heart of the present invention, the concrete steps of the method for constructing are as follows:

[0018] a. Preparation of cell culture medium

[0019] Select L-15 medium, add fetal bovine serum, human basic fibroblast growth factor and chondroitin sulfate to the medium, so that the volume of fetal bovine serum accounts for 20% of the final concentration of the total volume, human basic fibroblast The concentration of cell growth factor was 3 ng / ml and that of chondroitin sulfate was 0.5 μg / ml. The pH value is 7.0-7.4, and it is stored in a refrigerator at 4°C for later use;

[0020] b. Primary culture

[0021] First, soak fresh and live whitefish with 8 mg / L potassium permanganate for 10 min, then disinfect the fish as a whole, and then sterilize it again with 75% alcohol. In a 12-well plate, add PBS solution containing antibiotics to soak heart tissue, wherein: the concentration of penicillin is 200...

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Abstract

The invention relates to a construction method for a anabarilius grahami cardiac cell line and belongs to the technical field of freshwater organism cell culture. The method includes using anabarilius grahami heart tissue as a material, washing and shearing the heart tissue, using hyaluronidase and II type collagenase to perform a joint digestion to obtain free heart cells and loose tissue blocks, inoculating the free heart cells and the loose tissue blocks in a culture flask of 25 square centimeters, adding an L-15 culture solution containing fetal calf serum, human basic fibroblast growth factor and chondroitin sulfate in the culture flask, and placing the culture flask in a culture box of 28 DEG C to culture. The culture solution is replaced at an interval of three and half days, and when cells grow into monolayers, a trypsin digestive method is utilized to perform cell passage. The construction method for the schizothorax taliensis cardiac cell line has the advantages that the operation is simple and convenient, the cell growth state of the constructed anabarilius grahami cardiac cell line is good, the cells are fibroblast-like, more than 40 generations can be continuously achieved during the cell passage, the constructed anabarilius grahami cardiac cell line can be directly used for models of an environmental toxicology for studying environmental pollutants to perform pollution detection and safety evaluation, and the method is also suitable for the construction of cardiac cell lines of other fishes.

Description

technical field [0001] The invention relates to a method for constructing a heart cell line of the whitefish, which belongs to freshwater biological cell culture technology. Background technique [0002] Anabarilius grahami belongs to the order Cypriniforms, the family Cyprinidae, and the genus Anabarilius, commonly known as white fish. It is a rare fish species only produced in Fuxian Lake, Yunnan, and it is one of the four famous fish in Yunnan. The fish is tender and delicious, with soft spines and thin scales, and a delicious fragrance. This fish used to be the main fish species in Fuxian Lake. Over the years, due to the invasion of foreign fish species, the deterioration of water quality, and overfishing, it has been on the verge of extinction. Although since 1999, the Kunming Institute of Zoology, Chinese Academy of Sciences has successfully artificially propagated, protected and saved this rare fish species, and kept its output stable in recent years. However, the w...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/077
Inventor 潘晓赋王晓爱杨君兴陈小勇
Owner KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI
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