Construction method and ultralow temperature freezing and storing method of sinocyclocheilus grahami saccus olfactorius cell line

Abstract

The invention relates to a construction method and ultralow temperature freezing and storing method of a sinocyclocheilus grahami saccus olfactorius cell line, and belongs to the technical field of cell culture and ultralow temperature freezing and storing method of the biological cell of freshwater aquatic life. A sinocyclocheilus grahami saccus olfactorius tissue is used as a raw material to be cultured in an L-15 culture solution at the pH (Potential of Hydrogen) value of 7.0-7.2 and comprising fetal calf serum by means of a tissue explant and subculture is carried out by means of trypsin digestion. The method specifically comprises the preparation of a cell culture solution, primary culture and subculture. The construction method disclosed by the invention has the following beneficial effects: (1) primary culture takes short time, and cell mass is large and can be applied to chromosome analysis; (2) the constructed sinocyclocheilus grahami saccus olfactorius cell line is in fiber like morphology, can be subcultured continuously and directly applied to a research on biological characteristics, and satisfies the demands of the storage, the theoretical research and the application of sinocyclocheilus grahami germplasm resources; (3) the construction method is also applicable to the construction of saccus olfactorius cell lines of other fishes.

Description

Technical field: [0001] The invention relates to a method for establishing cell lines and cryopreservation at ultra-low temperature by using the olfactory sac tissue of the barb in Dianchi Lake, and belongs to the technical field of cell culture and cryopreservation of freshwater aquatic organisms. Background technique: [0002] Fish cell culture started in the 1960s, and has developed a relatively complete cell culture system including culture medium, antibiotics, primary culture and subculture methods. Up to now, more than 280 cell lines have been established. After more than 30 years of fish cell culture in my country, only more than 50 fish cell lines have been established, involving about 30 species, mainly marine species. From the above examples, it can be seen that a mature cell culture process is not a prerequisite for the successful establishment of a cell line. A mature technology process cannot make cell culture successful, and cell culture still needs to provide ...

AI Technical Summary

Problems solved by technology

Although the artificial propagation technology of golden-haired barb in Dianchi Lake has been successfully developed and released back to Dianchi Lake in a planned way, due to the limitations of the cultured fish itself, artificial breeding in the pond culture environment is likely to cause chromosomal abnormalities, such as polyploidy and aneuploidy, Short-term cell culture is required for karyotype analysis to test the quality of artificially propagated species and ensure the survival rate and reproduction rate of released individuals

For the fish such as the golden-haired catfish in Dianchi Lake, which is relatively small and has a small amount of blood, the traditional short-term blood culture cannot meet the needs of karyotype analysis; At around 22°C, the culture temperature of the cells is not high, and the temperature is the key factor affecting the growth rate of the cells. Therefore, the primary culture of the barb fin ray cell line in Dianchi Lake lasts for nearly two months, so it is necessary to find a fast Methods of Obtaining Cells to Satisfy the Short-Term Culture of Dianchichi goldenrod Cells for Karyotype Analysis

[0004] Through document retrieval, do not see identical report with the present invention