Schizosaccaromyce pombe biological genotoxicity screening system using recombinant fluorescence protein genes in yeast Schizosaccaromyce pombe and cultured human cells

摘要

The genes involved in DNA damage recovery and cytotoxicity related to cell and genetic toxicity, and their promoters, and promoters isolated from the genomic DNA were amplified by PCR amplification, and then their nucleotide sequences were verified and recombined into EGFP gene. After transforming the constructed recombinant gene into S.pombe and human cells, the molecular biological properties of the transformant were analyzed at the level of gene, transcript and protein, and the results of the gene expression of the transformants were in agreement with the expected , And dose dependence of DNA injury sources were identified at transcript level and protein level. A representative genotoxic source such as ultraviolet B and alkylating agent MMS was treated on the transformant having the characteristics, and the relative value of fluorescence, which is a response to the DNA injury source, was analyzed using a fluorometer measurement method. Increased DNA damage response to the increase and decrease of fluorescence over time of the normal culture medium after the toxicant treatment were determined and the strength of the useful promoters was determined. After establishing the standard reaction, various concentrations and time-course of the transformant to non-herbicidal toxins such as various cytotoxic agents such as ultraviolet C, MNNG, MNU, 4NQO, cisplatin, BPDE, 3MC, aflatoxin B and antibiotics The genetic toxicity screening system was validated only for the genotoxic source and the relative value of the maximum expression fluorescence to the untreated control group was determined. The response of this system to various environmental pollutants was compared, and the reaction of this system was also confirmed for some food residues. The yeast cell line and the activity of human cultured cells for these reactions were compared and the basis for industrial application was established.