Abstract
A functional gene coding for ribonuclease H was determined to apparently be essential for cell growth in E. coli. A strain was made with 2 copies of the rnh gene by lysogenizing an E. coli strain with a .lambda. phage bearing a copy of the rnh gene. Inactivation of 1 of the 2 copies of the rnh gene was accomplished by transformation with a linear DNA molecule that had the gene for chloramphenicol acetyltransferase inserted near the middle of the rnh gene. In recombinants that had an inactive gene replacing the normal chromosomal rnh gene, the .lambda.rnh prophage supplies an intact functional copy of the rnh gene. Curing the cells of the .lambda.rnh prophage left the cell with an inactive rnh gene and resulted in cell death. An intact functional rnh gene provided on a plasmid permits normal curing, and cured survivors were readily obtained. This technique is probably generally applicable for assessing the requirement for other E. coli genes.