Culture Control Parameters#
Parameters that control culture dilutions
name
The name of the culture. Example: E. coli or not inoculated.
description
A description of the culture. Example: replicate 1 or chemostat or turbidostat or morbidostat.
volume_vial
Volume of the vial in milliliters (ml). This is the liquid volume under the waste needle.
pump1_stock_drug_concentration
Concentration of the drug in the pump 1 stock bottle. Typically this should be 0 - pump1 should contain drug-free medium The unit of this parameter can be any unit of concentration (e.g., mg/L, Moles/L, mM, %, etc.). It is not specified but must be the same unit across all parameters involving drug dose.
pump2_stock_drug_concentration
Concentration of the drug in the pump 2 stock bottle. This is the highest concentration that can be achieved in the culture. The lowest nonzero concentration in the vial occurs when adding the smallest injectable volume of drug to the vial (limited by drop size ~0.05mL). The minimum inhibitory concentration of the culture should not be below the smallest injectable concentration (~1% of the stock concentration). The stock drug concentration should be at least the desired minimum inhibitory concentration (after adaptation)
dose_initialization
Initial dose added to the culture immediately when the experiment starts. -1 to disable. If this value is set to a positive number, the culture will get diluted as soon as the experiment starts. This is useful if you want to inoculate the culture into a vial with nonzero concentration of the drug. If enabled, this dose should be at least 1% of the pump2_stock_drug_concentration to ensure that at least a small droplet of the drug is added.
dilution_factor
Factor by which the population is reduced during dilution. For example, if the vial volume is 12 ml and the dilution factor is 1.6, the maximum volume of the culture during a dilution is 12 ml * 1.6 = 19.2 ml. A total volume of 7.2mL will be added at every dilution, but the ratio of pump1 and pump2 will be adjusted according to the stress increase parameters. To achieve chemostat or turbidostat conditions with minimal variation in OD, set the dilution factor to a small value (e.g., 1.1). Note that a higher dilution frequency limits the time window for OD-based growth rate estimation. A smaller dilution factor requires more device operations per generation. With every dilution the valve has to open and close, and an additional fixed waste volume is pumped to fill the waste tubing with air.
od_dilution_threshold
OD at which dilution occurs. -1 to disable OD triggered dilutions. This is useful for running replifactory in turbidostat mode.
delay_dilution_max_hours
Maximum time delay between dilutions. -1 to disable time-triggered dilutions. To achieve a dilution exactly every 2h, set this parameter to 2 and od_diultion_threshold to -1.
dilution_number_first_drug_addition
Dilution number at which dose_first_drug_addition is added. -1 to disable drug addition. The first initialization dilution is also counted; set this parameter to at least 2 if dose_initialization>0.
dose_first_drug_addition
Drug dose at first drug addition. -1 to disable drug addition. The dose is added to the culture at the dilution number specified by ‘dilution_number_first_drug_addition’. If enabled, this dose should be at least 1% of the pump2_stock_drug_concentration to ensure that at least a small droplet of the drug is added.
dose_increase_factor
Factor by which the dose is increased at stress increases. The new dose is calculated as: new_dose = old_dose * factor + amount. This parameter allows for a proportional increase in the drug dose based on the current dose.
dose_increase_amount
Amount by which the dose is increased at stress increases. The new dose is calculated as: new_dose = old_dose * factor + amount. This parameter ensures that the drug dose increases by a specified amount regardless of the current dose. It is particularly useful when the current dose is significantly lower than 1% of the pump2_stock_drug_concentration.
threshold_od_min_increase_stress
Minimum OD for stress increase to be allowed. Useful to prevent stress increases if the culture is not growing.
threshold_growth_rate_increase_stress
Minimum growth rate for stress increase to be allowed. Useful to prevent stress increases if the culture is not growing.
threshold_growth_rate_decrease_stress
If growth rate is below this value, stress is decreased by making one default dilution with pump1 only, lowering the drug concentration by ‘dilution_factor’. Period of consecutive stress decreases is controlled by ‘delay_dilution_max_hours’.
delay_stress_increase_min_generations
Minimum number of generations between stress increases. Useful to throttle the stress increase frequency.
postfill
Whether to fill up the vial after dilution (1 or 0). Modifies the order of pump operations in a dilution.
When postill is disabled, media is first added to the vial, then excess is pumped to the waste bottle. When postfill is enabled, excess media is first pumped to the waste bottle, then the vial is filled up.
Useful for high dilution factors.
For example, for a 1:15 dilution factor, elongate the waste needle so the dead volume is 1mL, set volume_vial to 1, dilution factor to 15 and postfill to 1.
Growth Rate and Doubling Time#
In the context of cell cultures, the growth rate is a measure of how quickly the cells in the culture replicate. The doubling time, on the other hand, is the amount of time it takes for the culture to double in size.
The relationship between growth rate (r) and doubling time (t) is given by the formula:
Where: - (log(2)) is the natural logarithm of 2, - t is the doubling time.
In other words, the growth rate is the reciprocal of the doubling time (scaled by the natural logarithm of 2), and vice versa. If you have a high growth rate, you’ll have a shorter doubling time, and if you have a long doubling time, your growth rate will be lower.
Let’s consider some examples with different growth rates:
For a growth rate of 0, the doubling time is infinitely long. This means the culture is not growing.
For a growth rate of 0.1, the doubling time is:
\[t = \log(2) / 0.1\]Which is approximately 6.93 hours.
For a growth rate of 0.5, the doubling time is:
\[t = \log(2) / 0.5\]Which is approximately 1.39 hours.
For a growth rate of 1, the doubling time is:
\[t = \log(2) / 1\]Which is approximately 0.69 hours, or about 41.4 minutes.