Short nucleotide sequence repetitions in DNA can provide selective benefits and also can be a source of genetic instability arising from deletions guided by pairing between misaligned strands. These findings raise the question of how the frequency of deletion mutations is influenced by the length of sequence repetitions and by the distance between them. An experimental approach to this question was presented by the heat-sensitive phenotype conferred by pcaG1102, a 30-bp deletion in one of the structural genes for Acinetobacter baylyi protocatechuate 3,4-dioxygenase, which is required for growth with quinate. The original pcaG1102 deletion appears to have been guided by pairing between slipped DNA strands from nearby repeated sequences in wild-type pcaG. Placement of an in-phase termination codon between the repeated sequences in pcaG prevents growth with quinate and permits selection of sequence-guided deletions that excise the codon and permit quinate to be used as a growth substrate at room temperature. Natural transformation facilitated introduction of 68 different variants of the wild-type repeat structure within pcaG into the A. baylyi chromosome, and the frequency of deletion between the repetitions was determined with a novel method, precision plating. The deletion frequency increases with repeat length, decreases with the distance between repeats, and requires a minimum amount of similarity to occur at measurable rates. Deletions occurred in a recA-deficient background. Their frequency was unaffected by deficiencies in mutS and was increased by inactivation of recG.