Fig. 4: Compacted state of chromosome in a TxTl system maintained by native condensin MukB.
From: Bacterial chromosome conformation and cell-free gene expression in synthetic 2D compartments
a A 3-D schematic of multiple E. coli chromosomes (typically >10) transplanted into a large diamond-shaped compartment, flanked by two flow channels used to introduce E. coli cells, various buffers, and the cell-free transcription-translation (TxTl) system. The compartment is closed by a glass coverslip to assemble the microfluidic chip. b The fluorescence signals of MukB-HT and RNAP (β’-HT) in separate experiments, each averaged over 4 chromosomes from two independent biological experiments in large compartments. The dashed colored lines show fits to mono-exponential decays (τMukB-HT = 6.89 ± 0.20 min and τRNAP = 2.63 ± 0.03 min) with immobile fractions (AMukB-HT = 0.27 ± 0.01 and ARNAP = 0.071 ± 0.002). c Four exemplary snapshots of chromosomes extracted from E. coli strains with (upper row showing double-color images) and without (lower row showing grayscale images) a mukB gene transplanted into large compartments. For both cases, the DNA was labeled with SYBR Green I (SG-I). MukB-HT was labeled with MaP655-Halo before cell lysis. d The estimated area of chromosomes with and without the mukB gene after transplantation into large compartments and ~25-minute incubation in TxTl and SG-I. The boxplot extends from the first quartile (25%) to the third quartile (75%) of the data, with a line at the median and a circle at the mean. The whiskers extend from the box to the farthest data point lying within 1.5-fold the inter-quartile range from the box. Individual chromosomes are shown as the smaller gray points (NWT = 67 and NΔmukB = 47), combined from two independent biological experiments. A two-sided Mann-Whitney U test was performed to compute the statistical significance between the two distributions. Source data is provided as a Source Data file.
