A PCR assay for rapid discrimination of sterile cytoplasm types in maize

A single-seed, multiplex polymerase chain reaction (PCR) protocol is presented to distinguish the three major types of cytoplasmic male sterility (CMS-T, -C, and -S) in maize (Zea mays L.). Three pairs of PCR primers were designed, corresponding to the chimeric regions of mitochondrial DNA sequences specific for the three types of CMS cytoplasm. The specificity of these primers was tested by means of the inbred line B37 with male-sterile (T, C. and S) and normal (N) cytoplasm as reference material. Subsequently, a single-seed PCR assay was developed for the determination of cytoplasm types in a single reaction and applied to a wide range of inbred lines with normal and sterile cytoplasm (T, C, and S) and with predominantly identified subtypes of the major sterile cytoplasm types. Our protocol proved to be a rapid, inexpensive, and reliable procedure for the molecular classification of male sterile maize cytoplasm to the three major categories T, C, and S.