Chromosomally encoded toxin–antitoxin (TA) systems are thought to result in growth arrest and eventual cell death upon exposure to environmental stress in E. coli. In the chromosome of cyanobacteria Synechocystis sp. PCC6803, the genetic organization of a 360 bp open reading frame (ORF), slr0664, and another small ORF of 256 bp, ssr1114, is similar to that of TA system. The predicted protein encoded by slr0664 is homologous to RelE, but neither homologue of ssr1114 nor ssr1114-encoding protein was found in TA system. To see whether slr0664 encodes a toxin protein, ssr1114 encodes an antitoxin, an expressing plasmid containing promoter Plac and PBAD, was constructed. In this construct, Both slr0664 and ssr1114 were controlled by Plac and PBAD, respectively. Expression of slr0664 in Escherichia coli results in the inhibition of bacterial growth, the expression of ssr1114 neutralize the toxicity of slr0664 expression. These results show that slr0664 is toxin gene and ssr1114 is antitoxin gene, both ssr1114 and slr0664 constitute a chromosomal TA system in Synechocystis sp. PCC6803.