| Sumario: | The lotus (<i>Nelumbo</i> Adans.) is an important aquatic plant with ornamental, medicinal and edible values and cultural connotations. It has single-, semi-double-, double- and thousand-petalled types of flower shape and is an ideal material for developmental research of flower doubling. The lotus is a basal eudicot species without a morphological difference between the sepals and petals and occupies a critical phylogenetic position in flowering plants. In order to investigate the genetic relationship between the sepals and petals in the lotus, the class <i>E</i> genes which affect sepal formation were focused on and analyzed. Here, <i>SEPALLATA 1</i>(<i>NnSEP1</i>) and its homologous genes <i>AGAMOUS-LIKE MADS-BOXAGL9</i> (<i>NnAGL9</i>) and <i>MADS-BOX TRANSCRIPTION FACTOR 6-like</i> (<i>NnMADS6-like</i>) of the class <i>E</i> gene family were isolated from the flower buds of the Asian lotus (<i>Nelumbo nucifera</i> Gaertn.). The protein structure, subcellular localization and expression patterns of these three genes were investigated. All three genes were verified to locate in the nucleus and had typical MADS-box characteristics. <i>NnSEP1</i> and <i>NnMADS6-like</i> were specifically expressed in the sepals, while <i>NnAGL9</i> was highly expressed in the petals, suggesting that different developmental mechanisms exist in the formation of the sepals and petals in the lotus. The significant functional differences between NnSEP1, NnMADS6-like and NnAGL9 were also confirmed by a yeast two-hybrid assay. These results expand our knowledge on the class <i>E</i> gene family in sepal formation and will benefit fundamental research on the development of floral organs in <i>Nelumbo.</i>
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