Iris laevigata is ideal for gardening and landscaping in northeast China because of its beautiful flowers and strong cold resistance. However, the short length of flowering time (2 days for individual flowers) greatly limits its applications. Molecular breeding and engineering hold high potential for producing I. laevigata of desirable flowering properties. A prerequisite is to identify and characterize key flowering control genes, the identity of which remains largely unknown in I. laevigata due to the lack of genome information. To fill this knowledge gap, we used sequencing data of the I. laevigata transcriptome to identify MADS-box gene-encoding transcription factors that have been shown to play key roles in developmental processes, including flowering. Our data revealed 41 putative MADS-box genes, which consisted of 8 type I (5 Mα and 3 Mβ, respectively) and 33 type II members (2 MIKC* and 31 MIKC^C, respectively). We then selected IlSEP3 and IlSVP for functional studies and found that both are localized to the nucleus and that they interact physically in vitro. Ectopic expression of IlSEP3 in Arabidopsis resulted in early flowering (32 days) compared to that of control plants (36 days), which could be mediated by modulating the expression of FT, SOC1, AP1, SVP, SPL3, VRN1, and GA20OX. By contrast, plants overexpressing IlSVP were phenotypically similar to that of wild type. Our functional validation of IlSEP3 was consistent with the notion that SEP3 promotes flowering in multiple plant species and indicated that IlSEP3 regulates flowering in I. laevigata. Taken together, this work provided a systematic identification of MADS-box genes in I. laevigata and demonstrated that the flowering time of I. laevigata can be genetically controlled by altering the expression of key MADS-box genes.

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