Calculate bacterial growth rates from plate reader kinetics data in R
Growth curves are commonly used in a microbiological studies to quantify the impact of various changes on bacterial growth rate (e.g., nutrient availability, gene expression). Modern microbial growth experiments can be arrayed in multi-well plates with absorbance measurements taken automatically by a platereader, which can result in thousands of individual datapoints.
GrowthRateR calculates the maximal growth rate of bacterial cell cultures in such growth experiments. This is done by log-transforming growth curves and applying a rolling regression with a shifting window, such that the maximum slope of any of the regressions is the maximal growth rate. This approach is advantageous for bacterial growth curves which do not nicely fit a sigmoidal curve (a first step for other tools, such as growthcurver).
GrowthRateR includes two functions:
growthrateR(): Calculate maximal growth rate of the cultures in each well then merges with experimental metadata. Includes option to automatically pre-process plate reader output file.growthrateR_single(): Plot the growth curve, regressions, and maximal growth rate for a single well.
Install directly from GitHub:
source("https://raw.github.com/kevinsblake/GrowthRateR/main/growthrate.R")Alternatively, can download and then install using the filepath:
source("dir1/dir2/growthrate.R")Load plate reader output data and platemap
platefile <- read_excel("dir1/dir2/platefile.xlsx")
platemap <- read_excel("dir1/dir2/platemap.xlsx")
Plate files must have a header containing: time, then each well (e.g., A1 to H12 for 96-well plates). The rows below contain the time of each measurement in hours (starting with zero) and the OD reading for each well. For example:
> head(EXP20220626.clean)
time A1 A2 A3 A4 A5 A6 A7 A8 A9 A10 A11 A12 B1 B2 B3 B4 B5 B6 B7
1 0.0000 0.098 0.094 0.102 0.102 0.099 0.098 0.098 0.095 0.096 0.095 0.097 0.100 0.095 0.129 0.122 0.119 0.124 0.122 0.124
2 0.0833 0.098 0.094 0.101 0.101 0.098 0.098 0.098 0.094 0.096 0.094 0.096 0.099 0.095 0.129 0.122 0.119 0.123 0.122 0.124
3 0.1667 0.098 0.094 0.101 0.100 0.098 0.098 0.097 0.094 0.096 0.094 0.095 0.099 0.094 0.129 0.123 0.119 0.124 0.123 0.124
4 0.2500 0.097 0.094 0.101 0.100 0.098 0.098 0.097 0.094 0.096 0.094 0.095 0.099 0.094 0.131 0.124 0.121 0.126 0.124 0.127
5 0.3333 0.097 0.094 0.101 0.100 0.098 0.098 0.097 0.094 0.096 0.094 0.095 0.099 0.094 0.134 0.127 0.123 0.128 0.126 0.128
6 0.4167 0.097 0.094 0.100 0.099 0.098 0.098 0.097 0.094 0.096 0.094 0.095 0.099 0.094 0.138 0.131 0.127 0.132 0.130 0.132If using the BioTek Gen5 program, editing to this format can be done automatically with the plate.clean=TRUE option.
IMPORTANT: check that the time column is numeric (e.g., 0, 10, 20) and that Excel hasn't auto-formatted it to 00:00:00 MM/DD/YYYY format.
Platemap file must have a header containing Well followed by any additional metadata useful for downstream analyses (e.g., strain, drug, concentration). Do not include units. For example:
Well code gene
1 A1 NA NA
2 A2 NA NA
3 A3 NA NA
4 A4 NA NA
5 A5 NA NA
6 A6 NA NA The Well column must have each well in the platereader file (e.g., A1 to H12 for 96-well plates), but rows with blank metadata columns (i.e. NA) will be removed.
Calculate maximal growth rate of the cultures in each well then merges with experimental metadata. Includes option to automatically pre-process plate reader output file.
growthrateR(platefile, platemap, timepoints=5, window=1, time.min=-Inf, time.max=Inf, plate.clean=FALSE)platefile Dataframe containing plate reader data
platemap Maps experimental metadata onto plate wells.
timepoints The frequency measurements were taken (in minutes; default = 5)
window The length of the window of the rolling regression (in hours; default = 1)
time.min The lowest timepoint value used. All measurements before this time are masked. (default = -Inf)
time.max The maximum timepoint value used. All measurements after this time are masked. (default = Inf)
plate.clean Cleans BioTek Gen 5 plate reader output platefile before processing. (default = FALSE)
# Generate growth rate df, with a window of 0.5 h, with platefile processing, ignoring measurements after 16h
df <- growthcurveR(EXP_platefile, EXP_platemap, window=0.5, time.max=16, plate.clean=TRUE)Plot the growth curve, regressions, and maximal growth rate for a single well.
growthrateeR_single(platefile, timepoints=5, window=1, time.min=-Inf, time.max=Inf, plate.clean=FALSE, well)platefile Dataframe containing plate reader data.
timepoints The frequency measurements were taken (in minutes; default = 5)
window The length of the window of the rolling regression (in hours; default = 1)
time.min The lowest timepoint value used. All measurements before this time are masked. (default = -Inf)
time.max The maximum timepoint value used. All measurements after this time are masked. (default = Inf)
plate.clean Cleans BioTek Gen 5 plate reader output platefile before processing. (default = FALSE)
well The well to be printed.
# Generate growth curve of well A1
plot <- growthcurveR_single(EXP_platefile, EXP_platemap, window=0.5, time.max=16, plate.clean=TRUE, well="A1")