Rfd1, a restorer to the Aegilops juvenalis cytoplasm, functions in fertility restoration of wheat cytoplasmic male sterility.

Cytoplasmic male sterility (CMS) is a crucial means for the utilization of heterosis, which is of great significance for improving the yield and quality of hybrids. Currently, fertility restoration has been extensively investigated in crops, but fertility restoration of CMS wheat with Aegilops juvenalis cytoplasm is poorly understood. Here, a backcross population BC1F1 derived from a cross between the male-sterile line Ju706A, its maintainer line 706B, and restorer line LK783 was used to map the Rfd1 locus by bulked segregant analysis and wheat 660K single nucleotide polymorphism genotyping. Ju706A displayed complete male sterility, and its fertility can be restored by LK783 with a pair of dominant genes Rfd1Rfd1. The locus was located to a 2.4 Mb region on chromosome 1BS by markers AX-174254104 and AX-111201011. Combined with transcriptomic analysis and quantitative real-time PCR assay, TraesCS1B02G197400LC, the most likely candidate gene for Rfd1, was found to encode a pectinesterase that was localized in the cell wall, and was highly expressed in fertile anthers. The silencing of Rfd1 resulted in decreased fertility, and heterogeneous expression of Rfd1 promoted pollen germination and affected vegetative growth. This implies that Rfd1 is required for anther or pollen development and male fertility in CMS wheat with Ae. juvenalis cytoplasm. Furthermore, a 7 bp deletion in Ju706A was employed to develop a specific marker, Xnwafu1, for molecular marker-assisted selection of restorers. This study provides a new understanding for exploring the fertility restoration mechanism of CMS.