In order to investigate the function of heat shock protein gene in Cucurbita moschata, we isolated the open reading frames (ORFs) of three HSP70 genes from C. moschata using transcriptome sequencing and RTPCR methods and named CmHSP701, CmHSP702 and CmHSP703. Their sequence length were 1 998, 1 941 and 2 118 bp, encoding 666, 647 and 706 amino acids, respectively. Protein sequence analysis showed that all three CmHSP70 were hydrophilic proteins with typical conserved domains of NBD and SBD. CmHSP701 contained signal peptide and transmembrane structure, and it was mainly located in endoplasmic reticulum. CmHSP702 and CmHSP703 did not contain signal peptide and transmembrane structure, and they were mainly located in cytoplasm and chloroplast, respectively. Homologous comparison and evolutionary analysis revealed that three CmHSP70 had the highest consistency with the HSP70 of Momordica charantia, Cucumis melo and Cucumis sativus, and the genetic distance was the closest. The qRTPCR analysis showed that three CmHSP70 could be induced in roots, stems, immature leaves and mature leaves by 42 ℃, and the expression level had obvious tissue specificity, with the highest in mature leaves. Under high temperature conditions, three CmHSP70 in mature leaves could be expressed in response to heat stress in a short time (0-2 h), especially CmHSP702. It is speculated that CmHSP702 may play an important regulatory role in the process of heat stress in C. moschata. The results provided a theoretical basis for further understanding the function of HSP70 and elucidating the thermotolerance mechanism of C. moschata.