Knocking out of the Mitochondrial Gene orf218 Using mitoTALECD Restores Cytoplasmic Male Sterility in Eggplant

In the alloplasmic lines of eggplant containing the cytoplasm of relative wild species, two types of cytoplasmic male sterility (CMS) are known: the pollen non-formation (PN) type and the anther indehiscent (AI) type. The gene responsible for PN-type CMS is presumed to be orf218, located in the mitochondrial genome. However, this presumption has been based on circumstantial evidence: (1) orf218 is present only in PN-CMS lines and their cytoplasmic donor, and (2) the amount of its transcripts differs between sterile and fertility-restored lines. To clarify the involvement of orf218 in PN-type CMS, we attempted to knockout orf218 using mitochondrial genome editing. We successfully modified the orf218 coding region using the mitoTALECD method, resulting in pollen formation in the mitochondrial genome-edited individuals. Since the pollen grains of these plants were functional and self-pollination was possible, we concluded that orf218 is the causative gene for PN-type CMS. However, morphological abnormalities were observed in the anthers of null-segregants obtained after the self-pollination of the initial transformant. This suggests that factors other than orf218 may also be involved in inhibiting normal stamen development in PN-type CMS eggplants.