Fractionation of Escherichia coli cell populations at different stages during growth transition to stationary phase

Abstract

Summary Cultures of Escherichia coli could be separated into more than 15 cell populations, each forming a discrete band after Percoll gradient centrifugation. The cell separation was found to result from the difference in buoyant density but not the size difference. The cell density increases upon transition from exponential growth to stationary phase. Exponential phase cultures formed at least five discrete bands with lower densities, whereas stationary phase cultures formed more than 10 bands with higher densities. Two molecular markers characterizing each cell population were identified: the functioning promoter species, as identified by measuring the expression of green fluorescent protein under the control of test promoters; and the expressed protein species, as monitored by quantitative immunoblotting. These findings together suggest that the growth phase‐coupled transition of E. coli phenotype is discontinuous.

The transcription regulator AllR senses both allantoin and glyoxylate and controls a set of genes for degradation and reutilization of purines

“…Promoter strength was determined as described previously (Makinoshima et al, 2002;Shimada et al, 2004). In brief, GFP was expressed under the control of a test promoter while red fluorescent protein (RFP) was under the control of a reference promoter.…”

“…In brief, GFP was expressed under the control of a test promoter while red fluorescent protein (RFP) was under the control of a reference promoter. The promoter assay vector pGRP carries two types of fluorescent protein genes, one for RFP under the control of reference promoter lacUV5 and the other for GFP under the control of a test promoter (Makinoshima et al, 2002;Shimada et al, 2004). The gcl promoter sequence upstream from the initiation codon was amplified by PCR using the genomic DNA from E. coli KP7600 as a template and a pair of primers, H026S (59-TGGGCAGAGATCTGCCACTGCATGCTTCCGGT-39) and H026T (59-TCATTTTATGCATTTTATTCCTACCCTA-39).…”

Isolation, Purification and Characterization of Nucleoids from <i>Synechococcus elongatus</i> PCC 7942

“…However, viable colony forming units was maximum at 120 h, and thereafter goes down slowly (Figure 1(B)). These results indicate that at the stationary phase of growth, heterogenous cell populations were not only observed in E. coli but also in Cyanobacteria [31] [32].…”

Proteomics of marine bacteria

“…Low and high salt concentrations 2-D PAGE and MALDI-MS [52] Heat shock 2-D DIGE and [ 35 S]methionine pulse-labelling combined with 2-D PAGE and MALDI-MS [54] Biofilm-specific proteins 2-D PAGE and MALDI-MS/MS or nano-LC-ESI-MS [79] Aerobic and anaerobic conditions 2-D PAGE and MALDI-MS [90] Cell isolation of symbiotic cells Cell density centrifugation to separate bacterial cell populations [14,15] Cell disruption of mixed bacterial assemblages Sonication with different lysis buffers [122] Image analysis Quantitative analysis of 2-D maps [37] Gel-image warping and quantitative analysis [38] 2 Marine genome sequencing projects…”

“…The separation of distinct fractions in a cell population has been demonstrated with the aid of density gradient centrifugation techniques using Nycodenz and Percoll, respectively. Makinoshima et al [122] were able to separate exponentially growing and stationary E. coli cell cultures into 15 distinct fractions using Percoll. Inoue et al [123] recently demonstrated the application of Percoll gradient centrifugation for the separation of various groups of bacteria in environmental water samples, which had been concentrated by filter techniques.…”