Growth Rates Made Easy

Abstract

In the 1960s-1980s, determination of bacterial growth rates was an important tool in microbial genetics, biochemistry, molecular biology, and microbial physiology. The exciting technical developments of the 1990s and the 2000s eclipsed that tool; as a result, many investigators today lack experience with growth rate measurements. Recently, investigators in a number of areas have started to use measurements of bacterial growth rates for a variety of purposes. Those measurements have been greatly facilitated by the availability of microwell plate readers that permit the simultaneous measurements on up to 384 different cultures. Only the exponential (logarithmic) portions of the resulting growth curves are useful for determining growth rates, and manual determination of that portion and calculation of growth rates can be tedious for high-throughput purposes. Here, we introduce the program GrowthRates that uses plate reader output files to automatically determine the exponential portion of the curve and to automatically calculate the growth rate, the maximum culture density, and the duration of the growth lag phase. GrowthRates is freely available for Macintosh, Windows, and Linux. We discuss the effects of culture volume, the classical bacterial growth curve, and the differences between determinations in rich media and minimal (mineral salts) media. This protocol covers calibration of the plate reader, growth of culture inocula for both rich and minimal media, and experimental setup. As a guide to reliability, we report typical day-to-day variation in growth rates and variation within experiments with respect to position of wells within the plates.

A Statistical Guide to the Design of Deep Mutational Scanning Experiments

“…A, C, D). This is due to the fact that calculating the log-ratios with respect to the sequencing depth D introduces a "saturation effect" such that the log-ratios are non-linear in t as if the mutants no longer grow exponentially (comparable to deceleration and saturation phase described in Hall et al 2014). In line with our observations (Figs.…”

Reconstructed Ancestral Enzymes Impose a Fitness Cost upon Modern Bacteria Despite Exhibiting Favourable Biochemical Properties

“…For growth curves in M9 minimal medium, LB starter cultures were used to inoculate M9 starter cultures which, after overnight incubation, were then used to inoculate growth curve flasks. OD 600nm readings of minimal medium cultures were taken every *45 min for 13 h. Growth rate constants during exponential growth were calculated using the programme GrowthRates (Hall et al 2014). The growth of DleuB (pUC19-ANC4) in minimal medium was also monitored over a 24-h period in a FLUOStar Optima microplate reader (BMG Labtech).…”

Experimental evolution and the dynamics of genomic mutation rate modifiers

“…Such studies should seek to address how common such pleiotropic effects are and whether their qualitative and quantitative effects on fitness can contribute to the maintenance of low wild-type mutation rates in natural populations. High-throughput growth rate screens (Blomberg, 2011;Hall et al, 2014) of mutant libraries could be conducted as a first approximation of fitness effects of mutation rate modifiers in model organisms such as yeast and E. coli; such growth rate measurements, however, may not capture all aspects of competitive fitness and may differ significantly in different propagation conditions (for example, MutS may be directly beneficial to bacteria in oxidizing environments: Torres-Barceló et al, 2013). Any comprehensive investigation of potential direct mutator fitness effects in asexual populations, moreover, will be complicated by the difficulty of separating direct fitness effects from indirect selective effects due to associated mutations.…”