Arsenic is highly effective in treating acute promyelocytic leukemia (APL), especially for relapsed patients. However, the treatment is highly affected by the resistance of the drug by patients, while the arsenic-resistance mechanism has not been well studied. A genome-wide screen was performed against a pool of 4 757 Saccharomyces cerevisiae mutants, each with one different gene individually deleted, to isolate genes that may mediate cellular resistance to arsenic. A one-step selection method was used. An aliquot of the pooled yeast library was plated on YPD agar plates supplemented with 3 mmol/L sodium arsenite. The genomic DNAs of the arsenic resistant strains were separately extracted, and amplified by PCR to get DNA fragments with UPTAG. The corresponding deleted genes were identified by comparing the PCR-amplified sequences with the UPTAG sequences from the Saccharomyces Genome Deletion Project. Mutations were identified in 104 genes/ORFs showing resistance to arsenic as compared to the wild type strain. To rule out the possibility that the resistant phenotype of these mutants is a result of arsenic-induced mutation during the screening process, the individual deletion strains from the mutant collection were picked up and tested individually for arsenic resistance using the spot assay. Of the 104 mutants identified in the screen, all exhibited significantly more resistance than the wild-type cells. Among the verified strains, 32 mutants turned out to have stronger phenotype that is resistant to 5 mmol/L arsenite. Five of the 32 mutants (FPS1, TMA20, UPF3, YAL066W, YOR309C) showed resistance to 7 mmol/L arsenite. The phenotype data were mapped onto the regulatory network. Bioinformatic studies of the genes revealed four neighborhoods, including mRNA catabolism, response to stress, histone acetylation, and protein synthesis and catabolism.