Cloning of controlling element Ac from waxy-m7

Unlike Ds, Ac elements are capable of autonomous transposition. Ac-induced mutations are not available at the Sh or Adh1 loci. There are, however, two Ac-induced mutations at the waxy locus. One of these, wx-m9, had been previously cloned by Fedoroff et al. The other mutant, wx-m7, differs from wx-m9 in several physiological respects. In homozygous condition, wx-m7 reverts to wild type earlier and more frequently than wx-m9. Other differences in the Ac action exhibited by these alleles are also observed (B. McClintock, Carnegie Inst. Wash. Yearbook 1963, 1964, 1965). We have cloned the Ac element from wx-m7 to examine these changes in phase at a molecular level.

A genomic library of wx-m7 was constructed in the lambda vector EMBL4. It was screened using a probe of the wild type locus. The clone contains a 4.3 kb insertion in the wx gene. The insertion is located about 2.5 kb towards the transcription start of the waxy gene relative to the insertion site of Ac in wx-m9. The Ac insertions in wx-m7 and wx-m9 are indistinguishable by restriction analysis and occur in the same orientation in both alleles. The waxy DNA outside of Ac shows some restriction polymorphism with the wild type allele. The double Ds structure hybridizes to both ends of the Ac element but not to internal fragments, suggesting that Ds is formed from an internal deletion of Ac.

Heteroduplex studies (kindly performed by I. Bertram of the Max-Planck-Institut fur Zuchtungsforschung, Koln-Vogelsang) indicate a 4.3 kb insertion relative to the wild type Wx gene as predicted by the restriction analysis. The insertion loop of Ac consistently shows a small stem structure of around 150 bp at its base. As the 11 bp inverted repeats found at the termini of Ds (see above) are insufficient to form such a stem structure, it will be interesting to see whether Ac from wx-m7 differs from Ds in sh-m5933 by sequence at the termini.

U. Behrens, N. Fedoroff,* A. Laird, M. Muller-Neumann, P. Starlinger and J. Yoder

*Carnegie Institution of Washington, Baltimore, MD

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