Pyruvate kinase (PK) is a key limited enzyme that regulates the glycolysis pathway, which play an important role in biological process including metabolism of basic substances in plants. Cyperus esculentus L. accumulating high levels of nutrients such as lipid/oil, starch, and sugar in tubers is an ideal model for investigating carbon metabolism and its regulatory mechanisms. A total of 7 CePK genes were identified by mining transcriptome data, including three plastidial CePKp (CePKpα, CePKpβ1 and CePKpβ2) and four cytosolic CePKc (CePKc1, CePKc2, CePKc3 and CePKc4). All CePK proteins encoded by CePK genes have the typical PK and PK_C domains. CePK proteins classified as the same type exhibit the similarity in sequence length, relative molecular weight, stability, and isoelectric point. CePKp proteins are hydrophilic proteins, and predicted to be located in the chloroplast. CePKc proteins are stable hydrophobic proteins, and predicted to be located in the cytoplasm. The tertiary structure prediction indicates that CePK proteins exist in the form of homologous tetramers. During the tuber germination and seedling establishing, the expression patterns of these CePK genes were significantly different, demonstrating that CePK genes may function differentially in regulating nutrient metabolism pathways in the tuber and seedling. The present data provide a new scientific basis for the in-depth analysis of the collaborative regulation mechanism underlying carbon flux and allocation and other biological processes in tuber germination and seedling establishing, as also benefiting breeding of high-yield and high-quality C. esculentus.