S1 and S2 DNAs are mitochondrial DNA (mtDNA) molecules found in male-sterile S cytoplasm (cms-S) of maize but not in fertile revertants with the nuclear background of inbred line M825 (Levings et al., Science 209:1021, 1983). Other revertants which were recovered from Wf9 lines of cms-S retained S1 and S2 DNAs (L.J. Escote et al., MNL 59:100, 1984; Ishige et al., Japan. J. Breed. 35:285, 1985). Three revertants designated J', G', R' were obtained as spontaneous mutants from J, G, and R sources of S cytoplasm in the Wf9 background. The results from mtDNA restriction endonuclease analyses of these revertants showed that the different restriction sites were present only in chromosomal mtDNA and not in S1 and S2 DNA.

Direct DNA-DNA hybridization experiments for these revertants were done to check the insertion of S1 and S2 DNA into the chromosomal mtDNA. Uncut mtDNA preparations extracted from revertants J', G', R' and sterile S cytoplasms were analyzed by agarose gel electrophoresis and Southern blots. S1 and S2 DNAs were extracted from another agarose gel by electroelution and purified by phenol-chloroform extraction. S2 DNA was nick-translated and used for the DNA-DNA hybridization probe. Autoradiographs of undigested mtDNAs showed that hybridization patterns of revertants and control cms-S were similar (Figure 1). The S2 DNA probe hybridized to S1 and S2 DNAs, and to S3, S4, and S5 DNAs as designated by Kemble and Mans (J. Mol. Appl. Genetics 2:161, 1983). Hybridization patterns of mtDNAs digested by XhoI were also similar to each other regardless of the different restriction fragment patterns of revertants (Figure 2).

These results indicate that the rearrangements of chromosomal mtDNA of revertants were not likely caused by the insertion of sequences homologous to S1 or S2 DNA.

Figure 1. An autoradiograph of Southern blot of the undigested mtDNA after hybridization with a probe of S2 DNA. Undigested mtDNAs isolated from S(1), J'(2), G'(3), R'(4), S1(5) and S2(6) were resolved by electrophoresis on 0.8% agarose gel.

Figure 2. An autoradiograph of Southern blot of the XhoI digest of mtDNA after hybridization with a probe of S2 DNA. MtDNAs isolated from S(1), J'(2), G'(3) and R'(4) were digested with XhoI and resolved by electrophoresis on 0.7% agarose gel.

Teruo Ishige, Fumio Takaiwa and Burle Gengenbach
 
 

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