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【Objective】Protein phosphatase (PP2C) is a type of protein phosphatase that exists in both animals and plants, In order to explore its important role in the ripening process of watermelon fruits.【Method】In this study, a PP2C gene ClPP2C3 was cloned in watermelon (Citrullus lanatus) by reverse transcription- polymerase chain reaction (RT-PCR), and its bioinformatics, expression pattern and subcellular localization analysis were conducted.【Result】The cDNA sequence length of ClPP2C3 in watermelon was 1 317 bp and it encoded 438 amino acids. The molecular weight of ClPP2C3 protein was 47.81 kD and its isoelectric point was 5.12. The ClPP2C3 protein contained one PP2C conserved domain and it had high homology with tomato SlPP2C3 in tomato and FaABI1 in strawberry that negatively regulated fruit ripening. ClPP2C3 in the watermelon was located in the nucleus. The gene quantitative expression analysis showed that the expression of ClPP2C3 in high-sugar-containing watermelons was significantly higher than that in low-sugar-containing watermelons. The 2 kb promoter (up 2 kb to ATG) activity of cultivated watermelon was significantly higher than that in ancestral watermelons, while there was no significant difference in the 1 kb promoter (up 1 kb to ATG) activity detected between these two species.【Conclusion】The SNPs in 1-2 kb promoter region might result in the difference of promoter activity in different varieties. [ABSTRACT FROM AUTHOR]