The RECl gene of Ustilago maydis exerts global control over the genetic system of this organism. Recombination, repair, mutageneisis, cell division, viability, and meiosis are all affected by mutation in the gene. Using a genomic library made in a replicating plasmid vector, we isolated a DNA fragment that complements the rec-l mutant. Preliminary evidence suggests that the RECl gene is contained on this DNA fragment. It is the subject of this proposal to characterize the RECl gene and investigate its role in controlling recombination. We plan several avenues of analysis. Using gene disruption techniques developed recently in our laboratory, we will definitively establish the identity of the cloned DNA fragment as the RECl gene. After subcloning and restriction mapping we will begin analysis of mRNA-DNA hybrids, nuclear run-on experiments, and primer extension mapping. These investigations will reveal the nature of the RECl gene transcript and shed light on factors regulating its level. The polypeptide product of the RECl gene will be established by hybrid selection and analyzed in vivo using antiserum raised against a fusion protein. The DNA sequence of the wild type gene will be determined and the lesions giving rise to the rec-l alleles will be identified. Finally, the gene will be separated from its natural promoter and placed under control of a different promoter sequence in order for us to study the biological consequences of heterologous control.
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