The dataset contains single-cell microscopy measurements of Escherichia coli bacteria grown in microfluic devices (mother machine design) for 72h, with M9 media at 37°C. Phase contrast images were obtained in 2 min intervals, while fluorescence images were taken every 10 min. Images were processed with DeLTA, a machine learning algorithm for automated cell segmentation. The data we provide is the result of this processing, structured to reproduce the figures of the linked manuscript. It contains information on single-cell growth (length, elongation rates), and fluorescence measurements indicate the mean intensity of GFP expressed under a plasmidial reporter of RpoS transcription or a constitutively expressed promoter (PA1).

Description of the data and file structure

Each data file (.rds) corresponds to a script file (.R), organized according to the figure numbering in the linked preprint manuscript. The correspondence between figure panels, datasets and associated scripts is provided in the Metadata file.

Data_Figure 1_Constitutive cells.rds: List of 2 elements, where the first contains mother cells and the second the daughters. Each element is a data frame of 32,399 rows and 8 columns, containing paired data for every cell division. Each row shows data of one generation. Columns include the cell lineage number (mat.well), time of birth (mat.time), mean fluorescence at birth (GFP), normalized fluorescence (normalized.GFP), elongation rates (elong.rates), promoter actitivy (promoter), normalized promoter actitivy (normalized.promoter), and standard deviation of intracellular fluorescence at birth (GFPsd).

Data_Figure 1_RpoS cells.rds: List of 2 elements, where the first contains mother cells and the second the daughters. Each element is a data frame of 21,748 rows and 8 columns, containing paired data for every cell division. Each row shows data of one generation. Columns include the cell lineage number (mat.well), time of birth (mat.time), mean fluorescence at birth (GFP), normalized fluorescence (normalized.GFP), elongation rates (elong.rates), promoter actitivy (promoter), normalized promoter actitivy (normalized.promoter), and standard deviation of intracellular fluorescence at birth (GFPsd).

Data_Figure 1_RpoS heatmap.rds: Matrix of mother cell RpoS mean fluorescence measurements over time. Each row is a different mother cell, and each column is the mean fluorescence at a given time point (10 min intervals).

Data_Figure 1_Constitutive heatmap.rds: Matrix of mother cell constitutive mean fluorescence measurements over time. Each row is a different mother cell, and each column is the mean fluorescence at a given time point (10 min intervals).

Data_Figure 2_Constitutive population.rds: List of 2 elements, where the first contains mother cells and the second the daughters. Each element is a data frame of 32,399 rows and 8 columns, containing paired data for every cell division. Each row shows data of one generation. Columns include time of birth (time), mean fluorescence at birth (fluorescence), promoter actitivy (promoter), normalized fluorescence (norm.fluorescence), normalized promoter actitivy (norm.promoter), elongation rates (elong.rates), and standard deviation of intracellular fluorescence at birth (cv).

Data_Figure 2_RpoS population.rds: List of 2 elements, where the first contains mother cells and the second the daughters. Each element is a data frame of 21,748 rows and 8 columns, containing paired data for every cell division. Each row shows data of one generation. Columns include time of birth (time), mean fluorescence at birth (fluorescence), promoter actitivy (promoter), normalized fluorescence (norm.fluorescence), normalized promoter actitivy (norm.promoter), elongation rates (elong.rates), and standard deviation of intracellular fluorescence at birth (cv).

Data_Figure 4A_ConstGFP_daughter transects_cropped.rds: Daughter cell data. Measurements taken at birth. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4A_ConstGFP_mother transects_cropped.rds: Mother cell data. Measurements taken at birth. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4B_RpoS_daughter transects_cropped.rds: Daughter cell data. Measurements taken at birth. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4B_RpoS_mother transects_cropped.rds: Mother cell data. Measurements taken at birth. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4C_ConstGFP_daughter transects_cropped.rds: Daughter cell data. Measurements taken at division. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4C_ConstGFP_mother transects_cropped.rds: Daughter cell data. Measurements taken at division. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4D_RpoS_daughter transects_cropped.rds: Daughter cell data. Measurements taken at division. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 4D_RpoS_mother transects_cropped.rds: Mother cell data. Measurements taken at division. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were cropped to exclude 10px off each side, then normalized into 20 longitudinal bins.

Data_Figure 5_RpoS_polar fluorescence.rds: List of 4 elements (1 = mother cells; 2 = daughter cells; 3 = daughter 2 cells; 4 = daughter 3 cells). Each element is a data frame, where each row is a measurement of a different generation. Data frames contain time (time.of.birth), old pole RpoS fluorescence at birth (op.bir), new pole fluorescence at birth (np.bir), old pole RpoS fluorescence at divison (op.div), new pole fluorescence at division (np.div), and a ratio op.bir/np.bir (bratio).

Data_Figure 5_ConstGFP_polar fluorescence.rds: List of 2 elements (1 = mother cells; 2 = daughter cells). Each element is a data frame, where each row is a measurement of a different generation. Data frames contain time (time.of.birth), old pole fluorescence at birth (op.bir), new pole fluorescence at birth (np.bir), old pole fluorescence at divison (op.div), new pole fluorescence at division (np.div), and a ratio op.bir/np.bir (bratio).

Data_Figure_6_old pole effect.rds: Data frame of mother cell data. Each row corresponds to a generation, and each column a measurement of cell physiology. It contains a cell lineage number (cell), length at birth (len.at.birth), mean fluorescence at birth (fluo1.at.birth), RpoS promoter activity (promoter), time (time.of.birth), and old pole length (oplen). The following measurements were recalculated by disregarding the old pole area of the cell: promoter2, fluo1.at.birth2, len.at.birth2. Paired daughter measurements are also provided for mean fluorescence (daughter.fluo1) and promoter activity (daughter.promoter).

Data_Figure S1_RpoS lineage.rds: Sample cell lineage, containing all cells measured in a sample growth well. The data is a large list (2 elements) of raw DeLTA output converted to a working R format. The first element is a list in which each element is a cell, measured from birth until the cell is lost or dead. The second element is a large array containing the raw cell segmentation information.

Data_Figure S1_RpoS mothers.rds: Data frame of mother cell data. Each row corresponds to a generation, and each column a measurement of cell physiology. It contains length at birth (len.at.birth), mean fluorescence at birth (fluo1.at.birth), RpoS promoter activity (promoter), and elongation rates (r).

Data_Figure S1_Constitutive lineage.rds: Sample cell lineage, containing all cells measured in a sample growth well. The data is a large list (2 elements) of raw DeLTA output converted to a working R format. The first element is a list in which each element is a cell, measured from birth until the cell is lost or dead. The second element is a large array containing the raw cell segmentation information.

Data_Figure S1_Constitutive mothers.rds: Data frame of mother cell data. Each row corresponds to a generation, and each column a measurement of cell physiology. It contains length at birth (len.at.birth), mean fluorescence at birth (fluo1.at.birth), constitutive promoter activity (promoter), and elongation rates (r).

Data_Figure S2_RpoS stationary.rds: List of 2 elements, where the first contains mother cells and the second the daughters. Each element is a data frame of 32,524 rows and 5 columns, containing paired data for every cell division. Each row shows data of one generation. Columns include time of birth in hours (time), time counted backwards from last maternal division (backwards.time), elongation rates (elong.rates), mean RpoS fluorescence at birth (fluorescence), and normalized fluorescence (norm.fluorescence).

Data_Figure S3.rds: List of 2 elements (mother and daughter measurements), each a data frame of fluorescence measurements over generations.

Data_Figure S6A_RpoS_mother transects.rds: Mother cell data. Measurements taken at birth. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were normalized into 20 longitudinal bins.

Data_Figure S6A_ConstGFP_mother transects.rds: Mother cell data. Measurements taken at birth. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were normalized into 20 longitudinal bins.

Data_Figure S6B_RpoS_daughter transects.rds: Daughter cell data. Measurements taken at birth. List of 109 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were normalized into 20 longitudinal bins.

Data_Figure S6B_ConstGFP_daughter transects.rds: Daughter cell data. Measurements taken at birth. List of 104 elements, where each element is a matrix of fluorescence measurements across cell lengths. Each list element stands for a generation, and each matrix row is a cell measured at that generation. Columns are fluorescence levels at a given intracellular localization, from old pole to new pole. Lengths were normalized into 20 longitudinal bins.

Data_Figure S5_cellct_mat_per_frame.rds: Matrix representing the number of cells present in a given growth well over time. Rows indicate a growth well (1 to 244) and columns the movie frame, in 10 min intervals (1 to 433).

Data_Figure S5_endtrans_list_per_frame.rds: List of 433 elements, each containing a matrix of RpoS fluorescence transects. Each element of the list stands for a movie frame, in 10 min intervals. In the matrices, each row contains fluorescence measurements along the axis of the last cell in the growth well.

Data_Figure S5_momtrans_list_per_frame.rds: List of 433 elements, each containing a matrix of RpoS fluorescence transects. Each element of the list stands for a movie frame, in 10 min intervals. In the matrices, each row contains fluorescence measurements along the axis of the mother cell.

Sharing/Access information

Links to other publicly accessible locations of the data:
Proenca, A. M., Tuğrul, M., Nath A., & Steiner, U. K. (2023). Declining old pole physiology gradually enhances gene expression asymmetry in bacteria. BioRxiv. https://doi.org/10.1101/2023.08.07.552301

Code/Software

This submission contains the following R files, organized as to replicate the figures of the linked manuscript with the provided datasets:

Figure 1.R
Figure 2.R
Figure 3.R
Figure 4.R
Figure 5.R
Figure 6.R
Figure S1.R
Figure S2.R
Figure S3.R
Figure S5.R
Figure S6.R
Table S2.R

Code was written in R v.4.1.2, using packages mgcv v.1.3-38, ggplot2 v.3.5.1, ggdensity v.1.0.0, and segmented v.1.4-0.