Construction from Mu d1 (lac Apr) lysogens of lambda bacteriophage bearing promoter-lac fusions: isolation of lambda ppheA-lac

Abstract

Bacteriophage Mu d1 (lac Aprr) was used to obtain strains of Escherichia coli K-12 in which the lac genes are expressed from the promoter of pheA, the structural gene for the enzyme chorismate mutase P-prephenate-dehydratase. A derivative of bacteriophage lambda which carries the pheA-lac fusion was prepared; the method used is generally applicable for the construction, from Mu dl lysogens, of specialized transducing lambda phage carrying the promoter-lac fusions. A restriction enzyme cleavage map of lambda ppheA-lac for the enzymes HindIII and PstI is presented.

Mu dX, a derivative of Mu d1 (lac Apr) which makes stable lacZ fusions at high temperature

“…In contrast, the Gal-Mu dl transductants often had a Lac phenotype that differed from the donor strain, indicating that these strains acquired an additional copy of Mu dl unlinked to gal during transduction. Transposition of Mu dl during Pl transduction has been observed previously (8). These results suggest that the Bx When Mu dX lysogens were selected for only their Apr phenotype, the Cmr phenotype was lost at a frequency of about 2%.…”

“…Second, Mu dl insertions are genetically unstable since secondary Mu-specific replicative transpositions occur at a relatively high frequency. This latter feature of Mu dl makes it impractical to use genetic selections based on the Mu dl operon fusion phenotype (8) and has necessitated the development of methods for stabilizing Mu dl fusions after their isolation (13).Previous studies have established that polar insertions in the Mu B gene (X mutations) prevent the expression of both B and kil and thus eliminate the temperature-sensitive growth phenotype of a Mu cts lysogen (2). Thus, we expected that an X mutation should eliminate the temperature sensitivity of a Mu dl lysogen.…”

“…Second, Mu dl insertions are genetically unstable since secondary Mu-specific replicative transpositions occur at a relatively high frequency. This latter feature of Mu dl makes it impractical to use genetic selections based on the Mu dl operon fusion phenotype (8) and has necessitated the development of methods for stabilizing Mu dl fusions after their isolation (13).…”

Identification of osmoresponsive genes in Escherichia coli: evidence for participation of potassium and proline transport systems in osmoregulation

“…Stabilization of Mu dl(Ap lac) lysogens. It is known that the Mu dl (Ap lac) prophage is proficient in transposition, a feature that can frustrate attempts to select for regulatory mutants from these lac fusion strains (14,36). To circumvent this problem, the two lac fusions that were characterized in some detail in this study were both stabilized so that Mu-mediated transposition could no longer occur.…”

Lambda placMu: a transposable derivative of bacteriophage lambda for creating lacZ protein fusions in a single step

“…However, the use of these phages does present some problems. First, the ability of Mu d(Ap lac) phages to transpose makes insertions of these phages genetically unstable because secondary insertions can occur at a relatively high frequency (14). This complicates certain genetic manipulations such as P1 transduction of a Mu d(Ap lac) prophage or selections for rare regulatory mutants based on the Lac phenotype of the fusion.…”