title: Dual fluorescent protein-based bioassay system for the detection of genotoxic chemical substances in Saccharomyces cerevisiae.
abstract: A new reporter system has been developed for quantifying the activity of potentially DNA-damaging substances in the yeast Saccharomyces cerevisiae. The system relies on two different reporter genes, yEGFP and DsRed-Express2, to screen for DNA-damaging chemicals. The yEGFP gene is fused to the test promoter of RNR2, whose measurable signal has a dose-dependent relationship with DNA damage. The gene encoding DsRed-Express2 is fused to a constitutive promoter of GPD, providing an internal control for normalizing cell numbers in the assay. The dual fluorescent protein assay system is performed by sequentially measuring the yEGFP and DsRed-Express2 fluorescent intensity of the same sample, with the results expressed as the ratio of yEGFP to DsRed-Express2 intensity (yEGFP/DsRed-Express2). The yeast fluorescent protein reporter assay was performed in 96-well microtiter plates in the presence of different concentrations of test substances, which were then characterized. The assay was very efficient, high-throughput, and amenable to full automation. Here, we demonstrate that this system can be used as a biosensor to assess the genotoxic potential of drugs and other chemical substances.
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.
authors.forename: Yixin
authors.initials: Y
authors.lastname: Lu
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.
authors.forename: Yongjie
authors.initials: Y
authors.lastname: Tian
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.
authors.forename: Ruikun
authors.initials: R
authors.lastname: Wang
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.
authors.forename: Qianqian
authors.initials: Q
authors.lastname: Wu
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: a Department of Preventive Medicine , Yangzhou Medical College, Yangzhou University , Yangzhou , Jiangsu , PR China and.|b Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Disease and Zoonoses , Yangzhou , PR China.
authors.forename: Xiangming
authors.initials: X
authors.lastname: Li
mesh_terms: D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D017930:Genes, Reporter / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics*; D008164:Luminescent Proteins / Q000235:genetics*; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012264:Ribonucleotide Reductases / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*
chemical_list: D008164:Luminescent Proteins; D009153:Mutagens; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C485184:enhanced green fluorescent protein; C413663:fluorescent protein 583; D049452:Green Fluorescent Proteins; C570014:RNR2 protein, S cerevisiae; D012264:Ribonucleotide Reductases
keywords: Chemical substances; DNA damage; DsRed-Express2 reporter; genotoxicity; yEGFP reporter; yeast