Rapid and precise mapping of the Escherichia coli release factor genes by two physical approaches

Abstract

The termination of protein synthesis in Escherichia coli requires two codon-specific factors termed RF1 and RF2. RF1 mediates UAA- and UAG-directed termination, while RF2 mediates UAA- and UGA-directed termination. The genes encoding these factors have been isolated and sequenced, and RF2 was found to be encoded in two separate reading frames. The map position of RF1 has been reported as 27 min on the E. coli chromosome, while the RF2 map position has not yet been identified. In this study, two new and independent methods for gene mapping, using pulsed field gel electrophoresis and an ordered bacteriophage library spanning the entire chromosome, were used to localize the map position of the RF2 gene. In addition, the location of the RF1 gene was more precisely defined. The RF2 gene is located at 62.3 min on the chromosome, while the RF1 gene is located at 26.7 min. This approach to mapping cloned genes promises to be a rapid and simple means for determining the gene order of the genome.