Non-radiate flowers are associated with truncation of the CYCLOIDEA gene in Scaevola aemula (Goodeniaceae)

Abstract

Floral morphology is a crucial trait that determines the commercial value of ornamental plants and pollinator preference. Scaevola aemula (Fan Flower) typically develops bilaterally symmetrical flowers, but a recessive mutant with a radially symmetrical flower (peloric) has been isolated from a breeding population. In the present study, we explored the molecular basis of the floral traits of S. aemula. We found a genetic-phenotypic association for the CYCLOIDEA (CYC) gene SaCYC2, which is known as a regulatory gene for floral symmetry in various plants. The peloric flower line had a nonsense mutation immediately downstream of the start codon of SaCYC2 causing a loss of function. RNA-sequencing analysis revealed three paralogous CYC genes in S. aemula, but only SaCYC2 was preferentially expressed in the dorsal side of the petals. Moreover, we developed a co-dominant cleaved amplified polymorphic sequence (CAPS) marker based on the nonsense mutation to verify floral traits, thereby accelerating the selection and breeding of Scaevola spp. Our findings support the conserved function of CYC genes that determine aspects of floral morphology in Asteraceae.

Keywords: CYCLOIDEA2; Scaevola aemula; Floral symmetry.

Fig. 1

Flower and whole plant of wild type, bilateral symmetrcal flower (A, B, C) and radially symmetric flower (peloric) (D, E, F). Dorsal slit is made during St3 to St4 (G). d, dorsal petal; l, lateral petal; v, ventral petal. (Scale bars: 1 cm.).

Fig. 2

A phylogenetic tree of CYCLOIDEA-related genes. A neighbor-joining tree is shown with bootstrap values (%) on the branches (1000 replicates). The scale bar indicates 0.05 amino acid substitutions per site.

Fig. 3

Gene structure of CYCLOIDEA2 of S. aemula. (A) Radially symmetric line #11,361 has nonsense mutation in SaCYC2 at the 99 bp position, C to A. Intermediate line #7952 is heterozygous for SaCYC2 and SaCYC2m, containing both the wild and mutant alleles. The frequencies of the wild and mutant alleles were visualized using IGV (Integrative Genomics Viewer). (B) CAPS marker for radial symmetry in S. aemula. The restriction enzyme Msel detects only mutated allele.

Fig. 4

The results of RT-qPCR of SaCYC1, SaCYC2, SaCYC3A and SaCYC3B on dorsal, lateral and ventral petal at St1 to St2 in S. aemula. Only SaCYC2 was expressed specifically in the dorsal region. Error bars of gene expression are ± 1 SD from three biological replicates. Different letters indicate statistically significant differences at p <0.05 according to Tukey’s HSD test.

Fig. 5

CAPS maker identifying radially symmetrical flower in S. aemula. (A) N/N, wild, BL shows band in approximately 151 bp, n/n, radially symmetrical line, RD shows bands in around 55 bp and 96 bp using this CAPS marker. N/n, hetero type shows bands both N/N and n/n. (B) Genotype using CAPS marker identified phenotype of BL, RD and IM. BL, bilateral symmetrical flower; RD, radially symmetrical flower; IM, intermediate flower.