期刊
TOXICOLOGY MECHANISMS AND METHODS
卷 27, 期 2, 页码 115-120出版社
TAYLOR & FRANCIS LTD
DOI: 10.1080/15376516.2016.1266540
关键词
RAD54 promoter; yeast cell; chemical mutagen; yEGFP; dose-effect relationship
类别
资金
- National Natural Science Foundation of China [81241099]
Two yeast enhanced green fluorescence protein (yEGFP) yeast reporter vectors, pR1558-yEGFP and pR406-yEGFP, which are regulated by two RAD54 promoters containing 406-bp and 1558-bp DNA sequences, respectively, were constructed using molecular biological techniques and transformed into yeast for the screening of genotoxins. The constructed biosensors were named W303-1A/R1558-yEGFP and W303-1A/R406-yEGFP. To quantify biosensor performance, both transformed yeast cells were exposed to multiple doses of genotoxins including methylmethane sulfonate (MMS; a DNA alkylating agent), 4-nitroquinoline-N-oxide (4-NQO; a DNA cleavage agent), 5-fluorouracil (5-Fu; an inhibitor of polymerases and topoisomerases) and colchicine and canavanine (affecting other biochemical activities). The yeast bioassay performance was analyzed using fluorescence-activated cell sorting (FACS) and Multi-Mode Reader in a 96-well black microplate. The observed W303-1A/R1558-yEGFP dose-effect relationship was more obvious and the maximum inductions were 5.96-fold (MMS), 2.19-fold (4-NQO) and 2.71-fold (5-Fu); the corresponding values for W303-1A/R406-yEGFP were 2.53-, 1.50- and 1.91-fold, respectively. It is suggested that it is best to select the entire RAD54 promoter when constructing recombinant yeast cells for screening mutagens.
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