A tissue culture-induced mtDNA mutation reverts during a second tissue culture period

Rearrangements of mitochondrial DNA (mtDNA) occur during culture and regeneration of plants from corn tissue cultures. One specific rearrangement recovered in plants from T cytoplasm cultures results in the loss of the largest BamHI restriction fragment and in the appearance of two new fragments (R.J. Kemble and D.R. Pring, 1982, Plant Infection, Springer-Verlag. p. 187-195; B.G. Gengenbach and D.R. Pring, 1982, Maize Biol. Res. p. 257-262).

We were interested in determining whether this rearrangement was stable during a second tissue culture period or whether reversions to the original T genome arrangement or to new arrangements would occur. The seventh seed generation of mutant line R2 (B.G. Gengenbach, et al., 1981, TAG 59:161-167) was the source of embryos for initiation of new tissue cultures. This R2 mutant line lacked the largest Bam fragment and had two new fragments compared with the standard T arrangement (Figure 1A and B). Plants were regenerated from R2 cultures and progeny lines were analyzed for mtDNA changes. Thirty-nine lines retained the same pattern as the R2 culture line (Figure 1C), but one line had a Bam restriction digest pattern similar to that of standard T (Figure 1D).

These results suggest either that the mtDNA organization of R2 could revert back to the nonmutant T pattern or that a heterogeneous mitochondrial population was maintained for seven sexual generations, from which assortment during culture resulted in an apparent homogeneous nonmutant T mtDNA population. Reversion of the mutant, perhaps via recombination within duplicate mtDNA regions, to the nonmutant arrangement seems more likely than the alternative of heterogeneity. More detailed molecular analyses in progress should resolve this issue.

Figure 1. Mitochondrial DNA from A188 inbred line versions of T cytoplasm (A); mutant R2 (B); a nonrevertant line representative of 39 lines regenerated from tissue cultures of mutant R2 (C); and the one revertant regenerated sister line (D). The mtDNA was digested with Bam HI and electrophoresed on 0.7% agarose gels for 30 hr.

Burle Gengenbach, Holly Jessen and Kathleen Storey

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