A new measure for gene prediction in eukaryotes is presented. The measure is based on the Discrete Fourier Transform (DFT) phase at a frequency of 1/3, computed for the four binary sequences for A, T, C, and G. Analysis of all the experimental genes of S. cerevisiae revealed distribution of the phase in a bell-like curve around a central value, in all four nucleotides, whereas the distribution of the phase in the noncoding regions was found to be close to uniform. Similar findings were obtained for other organisms. Several measures based on the phase property are proposed. The measures are computed by clockwise rotation of the vectors, obtained by DFT for each analysis frame, by an angle equal to the corresponding central value. In protein coding regions, this rotation is assumed to closely align all vectors in the complex plane, thereby amplifying the magnitude of the vector sum. In noncoding regions, this operation does not significantly change this magnitude. Computing the measures with one chromosome and applying them on sequences of others reveals improved performance compared with other algorithms that use the 1/3 frequency feature, especially in short exons. The phase property is also used to find the reading frame of the sequence.Gene prediction analysis, and specifically, the computational methods for finding the location of protein-coding regions in uncharacterized genomic DNA sequences, is one of the central issues in bioinformatics (Fickett 1996;Salzberg et al. 1998). For a given DNA sequence of an organism, in which the genes and other functional structures are not already known, it is very important to have an accurate and reliable tool for automatic annotation of the sequence: the number and location of genes, the location of exons and introns (in eukaryotes), and their exact boundaries (Claverie 1997). Therefore, along with standard molecular methods, many new methods for finding distinctive features of protein-coding regions have been proposed in the past two decades (see reviews by Fickett 1996;Claverie 1997;Mathé et al. 2002). These methods are based on different measures for discriminating between protein-coding regions and noncoding regions. Some of the measures are based on statistical regularities in genes or exons, which are not present in introns and intergenic sections, such as, for example, differences in codon usage (Staden and McLachlan 1982), hexamer counts (Claverie and