Cloning and mapping ofinfA, the gene for protein synthesis initiation factor IFI

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Abstract
The gene for translation initiation factor IF1, infA, has been identified by using two synthetic oligonucleotides to screen a Charon 30 library of Escherichia coli DNA. A recombinant lambda phage, C1921, was purified from a plaque positive for both probes. A 2.8 kb Bg/II fragment and a 2.0 kb HindIII fragment isolated from Cl92l were subcloned into the BamHI and HindIII sites of pBR322 to yield pTB7 and pTH2 respectively. Synthesis of IF1 in maxicells transformed with pTB7 or pTH2 indicates the presence of infA in both inserts. This was confirmed by DNA sequencing: a region was found that codes for a 8, 119 dalton protein with an amino acid sequence corresponding to IF1 The chromosomal location of infA was determined by mapping the closely linked beta-lactamase gene (Ampr) in pTB7 and pTH2. pTB7 and pTH2 were transformed into polA Hfr hosts, and integration of the plasmid by homologous recombination near infA was selected on the basis of ampicillin resistance. The site of integration was confirmed by Southern blot analysis of restriction nuclease digested wild type and transformed genomic DNA. The Ampr marker (and therefore infA) was mapped to about 20 minutes by Hfr interrupted matings and P1 transduction experiments. The structure and regulation of the infA operon currently are being investigated.