Abstract: This study started with four pectin methylesterase genes from Arabidopsis thaliana and two pectin methylesterase genes from the watermelon transcriptome data analysis of the West Muskmelon Molecular Breeding Research Center of Northeast Agricultural University. The method was used to identify the pectin methylesterase homologous genes of melon（Cucumis melo L.） and watermelon（Citrullus lanatus） in the Cucurbitaceae database, and perform their bioinformatics prediction analysis. The results showed that its structure and physicochemical properties were similar to those of Arabidopsis pectin methylesterase genes, and a phylogenetic tree was constructed for them.The tree results showed that the pectin methylesterase gene families of various species were different.The homology was high. In addition, this experiment also used the homologous cloning method to obtain the full-length of the watermelon Cla012554 gene and analyze the temporal and spatial expression of the gene. The sequencing results showed that the overall similarity between the cloned watermelon Cla012554 gene sequence and the comparison sequence was 95.62%, real-time fluorescent quantitative PCR analysis showed that Cla012554 gene was expressed in watermelon roots, stems,leaves and pulp, with the highest expression in the stem and the lowest expression in the root. At the same time, Cla012554 gene might also be involved in the ripening and softening of watermelon pulp.This study provided an important reference for further exploring the functions of PME genes in Cucurbitaceous crops.