Molecular aspects of genetic instability of an artificial 68 bp perfect palindrome in Escherichia coli

Abstract

An artificial 68 bp perfect palindrome carried on a plasmid (pAS807) is genetically unstable. An increase in the population of cells harbouring palindrome-deleted pAS807 derivatives (pAS807-V) is observed as the number of cell generations increases. The calculated frequency of palindrome excision events per cell generation and per plasmid replication round in Escherichia coli is 0.95 X 10(-4). Sequence analysis of eight independent isolates of palindrome-deleted molecules, reveals two symmetrical deletion types (three of type I and five of type II). The two types of pAS807-v molecules retain 19 bp of the original sequence of the 68 bp palindrome but differ in the content of the central 3 bp. The generation of the two deletion types is best explained by formation of intermediate cruciform structures. Following the fate of the palindrome in various bacterial mutants, we find that the excision events depend on functional polA1, polA(ex1), lig, texA343(recC343) and texA344(recB344) gene products. However, recB21 recC22 mutations do not affect palindrome excision.