Journal:

Article Title: Two New Early Bacteriophage T4 Genes, repEA and repEB , That Are Important for DNA Replication Initiated from Origin E

doi:

Figure Lengend Snippet: Bacteria, phages, and plasmids used in this study

Article Snippet: The E. coli strains, T4 phages, and plasmids used in this work are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, phage, or plasmid Relevant characteristic or genotype Reference or source E. coli B Wild type, sup 0 Our collection S/6 Smooth derivative of B; T6 resistant Our collection CR63 Wild type, supD Our collection BL21(DE3) F − sup 0 expresses T7 RNA polymerase 68 Novagen DH5α F − endA1 recA1 supE44 thi-1 φ80lacZΔM15 60 New England Biolabs Bacteriophages T4D Wild type Our collection T4D sip1 motA frameshift mutation Reference 28 and this work T4D N135 Gene 5 amber mutation 53 T4D B256 Gene 5 amber mutation 53 repEA1 Gene repEA amber mutation This work repEB1 Gene repEB ochre mutation This work repEA1-sip1 repEA motA double mutant This work repEB1-sip1 repEA motA double mutant This work Plasmids pET11a Ap r pBR322 origin; expression vector with T7 promoter Novagen pET11d Ap r pBR322 derivative; expression vector with T7 promoter Novagen pMal-c2 Ap r ColE1 origin; plasmid to construct fusions with E. coli malE New England Biolabs pGEM3 Ap r pBR322 derivative Promega pRV1 T4 repEA with His tag in pET11d This work pRV4 T4 repEB with His tag in pET11d This work pRV9 T4 repEB in pET11a This work pRV10 T4 repEA in pMal-c2 This work pRV11 T4 repEB in pMal-c2 This work pGL217 T4 Pst I- Xba I fragment, containing C-terminal segment of gene 5 , repEA , six iterons and early promoter in pGEM3 53 pGL233 Segment of T4 gene 5 in pGEM3 53 pGL501 T4 fragment containing beginning of gene 5 and repEB in pGEM3 53 Open in a separate window Bacteria, phages, and plasmids used in this study Construction of plasmids.

Techniques: Plasmid Preparation, Mutagenesis, Expressing, Construct

Journal:

Article Title: Two New Early Bacteriophage T4 Genes, repEA and repEB , That Are Important for DNA Replication Initiated from Origin E

doi:

Figure Lengend Snippet: Mapping, by primer extensions from primer 10, of the 5′ ends of early transcripts initiated from PE1. (A) Lane 1, RNA isolated from uninfected bacteria; lane 2, RNA isolated 4 min after infection of E. coli B with wild-type T4 at 30°C by using CsCl gradient purification (see Materials and Methods); lanes G, A, T, and C, sequencing reactions using the same primer on DNA of plasmid pGL217. (B) RNA isolated 4 min after infection of E. coli B with an RNAWiz kit (see Materials and Methods). Lane 1, wild-type T4; lane 2, sip1 (motA); lane 3, repEB1; lane 4, repEA1; lanes G, A, T, and C, sequencing reactions using the same primer on DNA of plasmid pGL217.

Article Snippet: The E. coli strains, T4 phages, and plasmids used in this work are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, phage, or plasmid Relevant characteristic or genotype Reference or source E. coli B Wild type, sup 0 Our collection S/6 Smooth derivative of B; T6 resistant Our collection CR63 Wild type, supD Our collection BL21(DE3) F − sup 0 expresses T7 RNA polymerase 68 Novagen DH5α F − endA1 recA1 supE44 thi-1 φ80lacZΔM15 60 New England Biolabs Bacteriophages T4D Wild type Our collection T4D sip1 motA frameshift mutation Reference 28 and this work T4D N135 Gene 5 amber mutation 53 T4D B256 Gene 5 amber mutation 53 repEA1 Gene repEA amber mutation This work repEB1 Gene repEB ochre mutation This work repEA1-sip1 repEA motA double mutant This work repEB1-sip1 repEA motA double mutant This work Plasmids pET11a Ap r pBR322 origin; expression vector with T7 promoter Novagen pET11d Ap r pBR322 derivative; expression vector with T7 promoter Novagen pMal-c2 Ap r ColE1 origin; plasmid to construct fusions with E. coli malE New England Biolabs pGEM3 Ap r pBR322 derivative Promega pRV1 T4 repEA with His tag in pET11d This work pRV4 T4 repEB with His tag in pET11d This work pRV9 T4 repEB in pET11a This work pRV10 T4 repEA in pMal-c2 This work pRV11 T4 repEB in pMal-c2 This work pGL217 T4 Pst I- Xba I fragment, containing C-terminal segment of gene 5 , repEA , six iterons and early promoter in pGEM3 53 pGL233 Segment of T4 gene 5 in pGEM3 53 pGL501 T4 fragment containing beginning of gene 5 and repEB in pGEM3 53 Open in a separate window Bacteria, phages, and plasmids used in this study Construction of plasmids.

Techniques: Isolation, Infection, Purification, Sequencing, Plasmid Preparation

Journal:

Article Title: Two New Early Bacteriophage T4 Genes, repEA and repEB , That Are Important for DNA Replication Initiated from Origin E

doi:

Figure Lengend Snippet: DNA synthesis measured by incorporation of [3H]thymidine into acid precipitable material (47) after infection of E. coli B with the indicated mutant and wild-type T4 strains at 30 (A) and 42°C (B). The reduced DNA synthesis of the single motA mutant compared with wild-type T4 is expected from the reduced expression of T4 replication and recombination genes in motA mutants (7, 11, 42, 66).

Article Snippet: The E. coli strains, T4 phages, and plasmids used in this work are listed in Table . table ft1 table-wrap mode="anchored" t5 TABLE 1 caption a7 Strain, phage, or plasmid Relevant characteristic or genotype Reference or source E. coli B Wild type, sup 0 Our collection S/6 Smooth derivative of B; T6 resistant Our collection CR63 Wild type, supD Our collection BL21(DE3) F − sup 0 expresses T7 RNA polymerase 68 Novagen DH5α F − endA1 recA1 supE44 thi-1 φ80lacZΔM15 60 New England Biolabs Bacteriophages T4D Wild type Our collection T4D sip1 motA frameshift mutation Reference 28 and this work T4D N135 Gene 5 amber mutation 53 T4D B256 Gene 5 amber mutation 53 repEA1 Gene repEA amber mutation This work repEB1 Gene repEB ochre mutation This work repEA1-sip1 repEA motA double mutant This work repEB1-sip1 repEA motA double mutant This work Plasmids pET11a Ap r pBR322 origin; expression vector with T7 promoter Novagen pET11d Ap r pBR322 derivative; expression vector with T7 promoter Novagen pMal-c2 Ap r ColE1 origin; plasmid to construct fusions with E. coli malE New England Biolabs pGEM3 Ap r pBR322 derivative Promega pRV1 T4 repEA with His tag in pET11d This work pRV4 T4 repEB with His tag in pET11d This work pRV9 T4 repEB in pET11a This work pRV10 T4 repEA in pMal-c2 This work pRV11 T4 repEB in pMal-c2 This work pGL217 T4 Pst I- Xba I fragment, containing C-terminal segment of gene 5 , repEA , six iterons and early promoter in pGEM3 53 pGL233 Segment of T4 gene 5 in pGEM3 53 pGL501 T4 fragment containing beginning of gene 5 and repEB in pGEM3 53 Open in a separate window Bacteria, phages, and plasmids used in this study Construction of plasmids.

Techniques: DNA Synthesis, Infection, Mutagenesis, Expressing