Project description:Study of the mechanisms of RecB mutant terminus DNA loss in Escherichia coli. FX158: WT MG1655 FX35: recB- FX37: ruvAB- FX51: matP- MIC18: recB- sbcD- sbcC- MIC20: recB- ruvAB- MIC24: matP- recB- MIC25: recA- recB- MIC31: sbcB- sbcD- MIC34: recA- recD- MIC40: linear chromosome MIC41: linear chromosome recB- MIC42: matP- ftsKC- MIC43: matP- ftsKC- recB- MIC48: recA- Cells were grown in M9 minimal medium supplemented with 0.4 % glucose to exponential phase (0.2 OD 650 nm). Chromosomal DNA was extracted using the Sigma GenElute bacterial genomic DNA kit. 5 μg of DNA were used to generate a genomic library according to Illumina's protocol. The libraries and the sequencing were performed by the High-throughput Sequencing facility of the I2BC (http://www.i2bc.paris-saclay.fr/spip.php?article399&lang=en, CNRS, Gif-sur-Yvette, France). Genomic DNA libraries were made with the ‘Nextera DNA library preparation kit’ (Illumina) following the manufacturer’s recommendations. Library quality was assessed on an Agilent Bioanalyzer 2100, using an Agilent High Sensitivity DNA Kit (Agilent technologies). Libraries were pooled in equimolar proportions. 75 bp single reads were generated on an Illumina MiSeq instrument, using a MiSeq Reagent kit V2 (500 cycles) (Illumina), with an expected depth of 217X. An in-lab written MATLAB-based script was used to perform marker frequency analysis. Reads were aligned on the Escherichia coli K12 MG1655 genome using BWA software. Data were normalized by dividing uniquely mapping sequence reads by the total number of reads. Enrichment of uniquely mapping sequence reads in 1 kb non-overlapping windows were calculated and plotted against the chromosomal coordinates.
Project description:Archer2011 - Genome-scale metabolic model of
Escherichia coli (iCA1273)
This model is described in the article:
The genome sequence of E.
coli W (ATCC 9637): comparative genome analysis and an improved
genome-scale reconstruction of E. coli.
Archer CT, Kim JF, Jeong H, Park JH,
Vickers CE, Lee SY, Nielsen LK.
BMC Genomics 2011; 12: 9
BACKGROUND: Escherichia coli is a model prokaryote, an
important pathogen, and a key organism for industrial
biotechnology. E. coli W (ATCC 9637), one of four strains
designated as safe for laboratory purposes, has not been
sequenced. E. coli W is a fast-growing strain and is the only
safe strain that can utilize sucrose as a carbon source.
Lifecycle analysis has demonstrated that sucrose from sugarcane
is a preferred carbon source for industrial bioprocesses.
RESULTS: We have sequenced and annotated the genome of E. coli
W. The chromosome is 4,900,968 bp and encodes 4,764 ORFs. Two
plasmids, pRK1 (102,536 bp) and pRK2 (5,360 bp), are also
present. W has unique features relative to other sequenced
laboratory strains (K-12, B and Crooks): it has a larger genome
and belongs to phylogroup B1 rather than A. W also grows on a
much broader range of carbon sources than does K-12. A
genome-scale reconstruction was developed and validated in
order to interrogate metabolic properties. CONCLUSIONS: The
genome of W is more similar to commensal and pathogenic B1
strains than phylogroup A strains, and therefore has greater
utility for comparative analyses with these strains. W should
therefore be the strain of choice, or 'type strain' for group
B1 comparative analyses. The genome annotation and tools
created here are expected to allow further utilization and
development of E. coli W as an industrial organism for
sucrose-based bioprocesses. Refinements in our E. coli
metabolic reconstruction allow it to more accurately define E.
coli metabolism relative to previous models.
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