The molecular characterization of mutations induced in Mutator lines at the Bronzel and Shrunken loci in maize

摘要

Restriction maps, of Shrunken (Sh) alleles in two standard maize lines, Q60 and B70, were determined by restriction endonuclease digestion and Southern blot hybridization. A comparison of these maps with previously determined restriction maps of Sh alleles from three different maize lines showed that restriction endonuclease target sites were highly conserved within the promoter and transcription unit. However, the region adjacent to the polyadenylation site contained a high degree of restriction fragment polymorphism. This comparison enabled us to determine that four sh mutations induced in Mutator lines were caused by DNA insertions into the 5[superscript]\u27 region of the Sh gene. In addition to an insertion at the 5[superscript]\u27 region of the gene, one mutant contained another insertion at the 3[superscript]\u27 terminus. Southern blot hybridization data showed that only two of the five insertions were structurally related to Mu 1 or Mu 1.7 elements, which suggests that transposable DNA, other than Mu 1 or Mu 1.7 is involved in Mutator activity. One of these insertions was molecularly cloned and, through Southern blot hybridization, shown to have no sequence homology with the entire Mu 1 element. Further analysis is required to determine what role, if any, these elements have in the Mutator system. The analysis of mutations induced in Mutator lines was continued by analyzing two bronzel mutations, bz1-Mum3 and bz1-Mum6. A 1.4 kb insertion structurally similar to Mu 1 caused the bz1-Mum3 mutation, whereas a smaller (960 bp) insertion caused the bz1-Mum6 mutation. Molecular cloning of this smaller insertion (Mu S) showed that it contained the terminal inverted repeats of Mu 1 but was missing a portion of the Mu 1 internal region, indicating that Mu S was a deletion derivative of Mu 1.