Kinesin superfamily of microtubule- based motor orchestrates a variety of cellular processes. Recent availability of mammalian genomes has enabled analyses of kinesins on the whole genome. Here we present a novel full-length kinesin prediction program (FKPP) for mammalian kinesin gene discovery based on a comparative genomics approach. Contrary to previous predictions of 94 kinesins, we identify a total of 134 potentially kinesin genes from mammalian genomes, including 45 from mouse, 45 from rat and 44 from human. In addition, FKPP synthesizes 25 potentially full-length mammalian kinesins based on the partial sequences in the database. Surprisingly, FKPP reveals that full-length human CENP-E contains 2701 aa rather than 2663 aa in the database. Experimentation using sequence specific antibody and cDNA sequencing of human CENP-E validates the accuracy of FKPP. Given the remarkable computing efficiency and accuracy of FKPP, we reclassify the mammalian kinesin superfamily. Since current databases contain many incomplete sequences, FKPP may provide a novel approach for molecular delineation of kinesins and other protein families.