title: Cloning the polB gene of Escherichia coli and identification of its product.
abstract: Using an in vivo mini-Mu cloning system, we have cloned the polB gene of Escherichia coli into the multicopy plasmid, pUC18. A chromosomal insert of 4.9 kilobases gave 30-40-fold overproduction of DNA polymerase II, and the cells containing the plasmid showed normal growth. The restriction pattern of the polB gene does not match that of either the polA gene or polC gene. Plasmid-directed protein synthesis demonstrates peptides of 99 and 82 kDa which are not expressed by derivative plasmids without DNA polymerase II activity. It appears from in situ gel assays and high performance liquid chromatography that 82- and 55-kDa proteins are derived from the 99-kDa protein by degradation, but all retain activity. DNA polymerase I or DNA polymerase III antibody does not inhibit the synthesis reaction of partially purified DNA polymerase II, but DNA polymerase II antibody does. By the criteria of restriction pattern of the polB gene, molecular weight of the protein, and antibody inhibition of reaction, DNA polymerase II can be demonstrated to be a distinct DNA polymerase.
authors.affiliation: Institute of Molecular Genetics, Baylor College of Medicine, Houston, Texas 77030.
authors.forename: H
authors.initials: H
authors.lastname: Chen
authors.forename: S K
authors.initials: SK
authors.lastname: Bryan
authors.forename: R E
authors.initials: RE
authors.lastname: Moses
mesh_terms: D000936:Antigen-Antibody Complex; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular / Q000379:methods; D004257:DNA Polymerase II / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005798:Genes, Bacterial; D008970:Molecular Weight; D010957:Plasmids; D015183:Restriction Mapping
chemical_list: D000936:Antigen-Antibody Complex; D004257:DNA Polymerase II