Six unique ORFs were characterized in tobacco plants with sua-CMS sterile cytoplasm, identifying the mtDNA basis for pollen sterility. sua-CMS (cytoplasmic male sterility), the most widely used sterile system in tobacco hybrids, is the only CMS type identified as having no negative effects on agronomic or quality traits in tobacco (Nicotiana tabacum) and as being fully male sterile. CMS is often associated with alterations of mitochondrial DNA (mtDNA), including novel chimeric open reading frames (ORFs), which result from rearrangement and recombination. Here, we obtained 34 mitochondrial ORFs in the sua-CMS line msZhongyan100 (sZY) by BLAST analysis. When we amplified these mitochondrial ORFs in seven tobacco CMS lines including sua-, glu-, rep-, rus-, tab1-, tab2-, and tab3-CMS types and in fertile tobacco, we found that six ORFs-orf82, orf103, orf115a, orf91, orf115b, and orf100-were located in three small regions (m-sr) of the mitochondrial genome of sZY and were unique to the sua-CMS line. We further amplified the m-sr fragments in three different backcross populations of the seven types of CMS, three F1 hybrids with sua-CMS sterile cytoplasm, two sua-CMS lines, and 284 fertile tobacco accessions. The ORFs were specific to plants with the sua-CMS background. All six unique ORFs were chimeric and had no homology with the mitochondrial genomes of fertile tobacco. Transcript analysis revealed that the ORFs were highly expressed in the anthers and floral buds of sZY. These six ORFs were specific to sua-CMS and could be used as molecular markers to identify sua-CMS lines, which is useful for improving breeding for heterosis in tobacco.
Keywords: Mitochondrial ORF; Molecular markers; Tobacco; Transcript analysis; sua-CMS.