To investigate the effect of phosphorylation on the transcription activity of p53, ten phosphorylation mutants were constructed covering all the identified phosphorylation sites of rat p53. These included mutants of two casein kinase I sites (Ser6 and Ser9), two DNA-PK sites (Ser15 and Ser39), a p34cdc2 site (Ser313), the adjacent Ser312 and a casein kinase II site (Ser390). Two double phosphorylation mutants (Ser4, 6 and Ser15, 390) and one triple phosphorylation mutant (Ser4, 6 and 15) were also constructed. The transcription activity of all the p53 phosphorylation mutants was tested by transfection into two different types of cells, Saos-2 cells and p53(-/-) fibroblasts derived from p53 knock out mice, which both lack endogenouse p53. Surprisingly, all the p53 phosphorylation mutants retain transcription activity and the seven mutants tested can also suppress cell growth.