Abstract: To explore the genome size and characteristics of Nuphar pumila, this study selected 3 N. pumila samples, numbered KMPPC01, KMPPC02, and KMPPC03 for flow cytometry detection. The KMPPC01 was selected for high-throughput sequencing. Combined with bioinformatics methods such as k-mer analysis, preliminary research was conducted on the genome of N. pumila. The results are as follows: The genome size of N. pumila was determined to be approximately 2187.12 Mb by flow cytometry and the converted C value was approximately 1.12 pg; the raw data obtained by high-throughput sequencing was 111.76 Gb, and the clean data obtained after filtering was 108.39 Gb; K-mer analysis showed that the genome size of N. pumila was 2094.71 Mb, the GC content was 38.20%, the heterozygosity was 1.89%, the proportion of repeated sequences was 62.63%. In summary, the genome of N. pumila is a highly heterozygous and highly repetitive complex genome with a size of approximately 2094.71–2187.12 Mb.