Category:Strain:BW25113
See Help:Strains for more information about strains in EcoliWiki.
| Name |
BW25113 |
|---|---|
| Synonyms | |
| Genotype |
F- DE(araD-araB)567 lacZ4787(del)::rrnB-3 LAM- rph-1 DE(rhaD-rhaB)568 hsdR514 |
| Phenotype | |
| Plasmids | |
| Phage | |
| Sources | |
| Construction |
see description below |
| References | |
Notes
BW25113 is a derivative of the F-, λ-, E. coli K-12 strain BD792 (CGSC6159)[1], which is a derivative of E. coli K-12 W1485. The steps in the construction of BW25113 are outlined below.
i) W1485 ([CGSC# 5024]) genotype: F+ LAM- rpoS396(Am) rph-1 -->
ii) BD792 ([CGSC# 6159]) genotype: F- LAM- rpoS396(Am) rph-1. This is an F- derivative of W1485, cured with acridine orange. -->
iii) An rpoS+ derivative of BD792 was made in two steps. BD792 "was first made tetracycline resistant and kanamycin resistant by using P1kc grown on ZK1001 (cysC95::Tn10 rpoS::kan; from R. Kolter). A resultant Cys- transductant was then made cysteine independent and kanamycin sensitive by using P1kc grown on MG1655. E. coli K-12 strains BW25113, BW25141, BW25142, and BW25695 are descendants of BD792 derivatives that were made rpoS+ ([3])." -->
The remaining steps in the construction of BW25113 are described in Supplementary Table 1 of Baba et al. ([2]). At each step, mutant alleles were constructed in vitro and recombined into the genome using oriRR6Kgamma plasmids, which cannot replicate in a strain that lacks the R6K pir gene ([4]). In the absence of selection of the antibiotic resistance marker carried on the plasmid, homologous recombination can occur that will either restore the original genotype or result in allele replacement.
<TableEdit> Strain Genotype Plasmid used Reference </TableEdit>
iv) BW21597 genotype: DEphoR574 rph-1 [5]
v) BW21652 genotype: DEphoR574 DE(creABCD)154 rph-1 [4]
vi) BW23186 genotype: DEphoR574 DE(creABCD)154 DE(araBAD)567 rph-1 [6]
vii)
BW25113 was used to make the Keio Collection of single-gene knockouts. [2]
[1] Available from the Coli Genetics Stock Center: CGSC#7636
The genotype for BW25113 in Datsenko et al. [1] said the strain is lacIQ, but it was later shown to be lacI+ Baba et al. (Supplementary Table 1)[2].
According to the CGSC, lacZ4787(del) has 4 tandem copies of the rrnB transcriptional terminator inserted by gene replacement into the region extending from near the SacII site near the N-terminus of lacZ through the promoter.
References
See Help:References for how to manage references in EcoliWiki.
- ↑ 1.0 1.1 1.2 1.3 Datsenko, KA & Wanner, BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl. Acad. Sci. U.S.A. 97 6640-5 PubMed
- ↑ 2.0 2.1 2.2 2.3 Baba, T et al. (2006) Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection. Mol. Syst. Biol. 2 2006.0008 PubMed
- ↑ Haldimann, A & Wanner, BL (2001) Conditional-replication, integration, excision, and retrieval plasmid-host systems for gene structure-function studies of bacteria. J. Bacteriol. 183 6384-93 PubMed
- ↑ 4.0 4.1 Metcalf, WW et al. (1996) Conditionally replicative and conjugative plasmids carrying lacZ alpha for cloning, mutagenesis, and allele replacement in bacteria. Plasmid 35 1-13 PubMed
- ↑ Haldimann, A et al. (1997) Transcriptional regulation of the Enterococcus faecium BM4147 vancomycin resistance gene cluster by the VanS-VanR two-component regulatory system in Escherichia coli K-12. J. Bacteriol. 179 5903-13 PubMed
- ↑ Haldimann, A et al. (1998) Use of new methods for construction of tightly regulated arabinose and rhamnose promoter fusions in studies of the Escherichia coli phosphate regulon. J. Bacteriol. 180 1277-86 PubMed