The use of transposable elements as tags allows the isolation of genes whose products are unknown. Since En/Spm and Mu-1-induced al mutants are available we took advantage of the Mu-1 DNA and the recently cloned En-element (see accompanying report by Berndtgen et al.) to clone the A1 gene.
Genomic libraries were prepared from both maize stocks, containing either the En-induced a1-m(papu) mutation (Peterson, P.A. Theor. Appl. Genet. 40:367377, 1970) or Robertson's a1-Mum2 mutant (Robertson, D.S. Mutat. Res. 51:21-28, 1978. These libraries were then screened with En1 and Mu1-specific probes, respectively. However, since En1 and Mu1 are present in many copies in the maize genome, cross screening of the positives from each of the two libraries facilitated the identification of the clones containing the a1 gene. Using the a1 part of the two mutant clones the gene could be isolated from a wildtype line (the clone is available and has already been distributed).
A similar strategy to that described above was used in the isolation of the C2 locus. One of the mutant lines used is the Spm(En)-induced c2 mutant c2-m1 (the autonomous element is present at the C2 locus (McClintock, B. Carnegie Inst. Year Book 65:568-578, 1967). Since the screening of the En-induced a1 mutant library resulted in the isolation of only one clone identical to the autonomous En element present in wx-844, we assumed that the same would be true for the c2-m1 line. The analysis of this library, however, showed the presence of several copies of an element very similar to the autonomous En element in size and sequence. This 8kb-long element is identical to the autonomous En element over a 6kb internal region but differs partially in the terminal sequences.
The presence of this element, which is structurally very similar to an autonomous En element, causes problems in quickly screening the library, because the analysis of each positive clone has to be done very carefully. This element is also present in a Spm-induced c2 mutant line (c2-m2) containing the receptor (I) part of the element at the locus. The screening for a clone containing the C2 gene is still going on.
Catherine O'Reilly, Nancy S. Shepherd, Andy Pereira, Zsuzsanna Schwarz-Sommer, Ulla Niesbach, Ulrike Weydemann, Udo Wienand, Donald S. Robertson1, Peter A. Peterson2 and Heinz Saedler
1Dept. of Genetics, 2Dept. of Agronomy, Iowa State University, Ames, Iowa
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