Abstract
Two forms of male-sterile cytoplasm, designated nap and pol, are found in the oilseed rape species, Brassica napus. The nap cytoplasm is observed in most B. napus varieties, and it confers male sterility on a limited number of cultivars that lack the corresponding restorer gene, Rfn. In the present study, using linkage analysis in combination with 5652 BC1 progeny derived from a cross between a nap cytoplasmic male sterility (CMS) line 181A and a restorer line H5, we delimited the Rfn gene to a 10.5 kb region on chromosome A09, which contained three putative ORFs. Complementation by transformation rescue revealed that the introduction of ORF2, which encodes a pentatricopeptide repeat (PPR) protein, resulted in the recovery of fertility of nap CMS plants. Expression analysis suggested that the Rfn was highly expressed in flower buds and it was preferentially expressed in the tapetum and meiocytes during anther development. Further RNA gel blots and immunodetection suggested that the Rfn gene may play a complicated role in restoring the nap CMS. Our work laid the foundation for dissecting the molecular basis of CMS fertility restoration and the nuclear-mitochondrial interactions in CMS/Rf systems.
Highlights
Cytoplasmic male sterility (CMS), a maternally inherited trait that fails to produce functional pollen, has been observed in >150 higher plant species
Comparing the putative promoter region (~2.0 kb) and the full-length CDS of Rfn from H5 with that of the rfn allele from 181A, we found that only five single nucleotide polymorphisms (SNPs) were detected in the putative promoter region, while rich sequence variations including 184 SNPs were identified in the coding sequence
A previous study indicated that the restorer genes Rfn for nap cytoplasm and Rfp for pol cytoplasm represent different alleles or haplotypes of a single nuclear genetic locus (Li et al, 1998)
Summary
Cytoplasmic male sterility (CMS), a maternally inherited trait that fails to produce functional pollen, has been observed in >150 higher plant species. The male sterility phenotype can be recovered by dominant nuclear genes, termed restorer of fertility (Rf) genes, which can reduce the accumulation of CMS-associated RNAs or proteins in F1 hybrids. The nap cytoplasm is the most prevalent cytoplasm in B. napus germplasm, but it confers male sterility only in a limited number of cultivars that lack the corresponding fertility restorer gene, Rfn. In the absence of the nuclear restorer gene Rfn, plants carrying the nap cytoplasm display male sterility when grown under relatively low temperature (22/16 °C; 16/8 h), while the sterility can be gradually repaired or even completely reversed with an increase of temperature (Fan and Stefansson, 1986). The pol CMS is highly stable in most cases; it can be widely used for commercial hybrid seed production (Fan and Stefansson, 1986; Zhou and Fu, 2007)
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