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Updated: Aug 14, 2026

Mutagenesis and Functional Selection Protocols for Directed Evolution of Proteins in E. coli
Published on: March 16, 2011
New developments on the generation of mutations in Escherichia coli lysogens
1Department of Microbiology, Stritch School of Medicine, Loyola University, Maywood, Illinois.
Abstract:
Through the lytic process, P1 is a bacteriophage or virion capable at very low frequency of carrying out generalized transduction between strains of Escherichia coli. When P1 is not involved in lytic functions, it exists as a prophage in the form of circular DNA molecule, persisting in an extra-chromosomal or plasmid state, and not integrating into the host chromosome. E. coli carrying such plasmids have been referred to as lysogens. Earlier research dealt with P1 plasmids carrying drug-resistant factors. Up to the present study, P1 plasmids of any type were not known to have any mutagenic effect on the E. coli chromosome (genome), nor was it known that generalized transduction can be associated with mutagenicity. From P1 plasmids carrying a chloramphenicol resistance-factor, mutant plasmids of a particular type were isolated in the present study. P1 plasmids of this type carried a mutant factor which greatly impaired the capacity of phage derived from such plasmids, upon the completion of the lytic cycle, to lysogenize recA E. coli through phage promoted recombination. The plasmid of this mutant type is referred to as P1CMrec, and lysogens carrying such are referred to as P1CMrec lysogens. This paper describes the history of these P1CMrec lysogens and the genetic mutability within the E. coli chromosome of such P1CMrec lysogens, and the relationship of such genetic mutability (instability) to the incorporation of virion-DNA, following the absorption of P1 phage by these lysogens. As illustrated in the paper, a mutagenic effect was generated within the E. coli chromosome of P1CMrec lysogens by means of the P1CMrec plasmid. Furthermore, this mutagenic effect was found to be greatly, non-locally, and uniformly enhanced as a consequence of P1 virion incorporation by, or likely generalized transduction of, such lysogens. More specifically, the plasmid of this mutant type (P1CMrec) is responsible for the creation of a wide range of genetic mutabilities (instabilities) of differing degree within the E. coli genome (not carrying recA), some mutabilities being very high upon extended incubation. The P1CMrec plasmid was also involved in the creation of new mutant genes within the E. coli genome (not carrying recA), some mutabilities being very high upon extended incubation. The P1CMrec plasmid was also involved in the creation of new mutant genes within the E. coli chromosome, some of which manifested high mutability.(ABSTRACT TRUNCATED AT 400 WORDS)
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