Haplotype-Resolved, Chromosome-Level Assembly of White Clover (Trifolium repens L., Fabaceae).

White clover (Trifolium repens L.; Fabaceae) is an important forage and cover crop in agricultural pastures around the world and is increasingly used in evolutionary ecology and genetics to understand the genetic basis of adaptation. Historically, improvements in white clover breeding practices and assessments of genetic variation in nature have been hampered by a lack of high-quality genomic resources for this species, owing in part to its high heterozygosity and allotetraploid hybrid origin. Here, we use PacBio HiFi and chromosome conformation capture (Omni-C) technologies to generate a chromosome-level, haplotype-resolved genome assembly for white clover totaling 998 Mbp (scaffold N50 = 59.3 Mbp) and 1 Gbp (scaffold N50 = 58.6 Mbp) for haplotypes 1 and 2, respectively, with each haplotype arranged into 16 chromosomes (8 per subgenome). We additionally provide a functionally annotated haploid mapping assembly (968 Mbp, scaffold N50 = 59.9 Mbp), which drastically improves on the existing reference assembly in both contiguity and assembly accuracy. We annotated 78,174 protein-coding genes, resulting in protein BUSCO completeness scores of 99.6% and 99.3% against the embryophyta_odb10 and fabales_odb10 lineage datasets, respectively.

Transcriptomic Analysis Suggests Auxin Regulation in Dorsal-Ventral Petal Asymmetry of Wild Progenitor Sinningia speciosa.

The establishment of dorsal-ventral (DV) petal asymmetry is accompanied by differential growth of DV petal size, shape, and color differences, which enhance ornamental values. Genes involved in flower symmetry in Sinningia speciosa have been identified as CYCLOIDEA (SsCYC), but which gene regulatory network (GRN) is associated with SsCYC to establish DV petal asymmetry is still unknown. To uncover the GRN of DV petal asymmetry, we identified 630 DV differentially expressed genes (DV-DEGs) from the RNA-Seq of dorsal and ventral petals in the wild progenitor, S. speciosa 'ES'. Validated by qRT-PCR, genes in the auxin signaling transduction pathway, SsCYC, and a major regulator of anthocyanin biosynthesis were upregulated in dorsal petals. These genes correlated with a higher endogenous auxin level in dorsal petals, with longer tube length growth through cell expansion and a purple dorsal color. Over-expression of SsCYC in Nicotiana reduced petal size by regulating cell growth, suggesting that SsCYC also controls cell expansion. This suggests that auxin and SsCYC both regulate DV petal asymmetry. Transiently over-expressed SsCYC, however, could not activate most major auxin signaling genes, suggesting that SsCYC may not trigger auxin regulation. Whether auxin can activate SsCYC or whether they act independently to regulate DV petal asymmetry remains to be explored in the future.