Growth curve for anaerobic bacteria

[catherineandrews]

Hi, I have very little micro experience and would like some advice please! I need to do a growth curve by measuring OD600 at 30min intervals. My bacteria is anaerobic and I usually use the anaerocults in a gas jar to produce an anerobic enviroment. However, as these take 20mins to reach anaerobic conditions I am thinking that this will affect the growth of my bacteria if I am opening the jar every 30mins. I could just do the OD every 60mins but every 30mins would be best. I also could use an anaerobic chamber but I think this is around 18% CO2, while the anaerocults are 5% CO2 (maybe incorrect?!). Would the difference in CO2 have an effect on growth or is it ok once the environment is anaerobic whatever the % of CO2? Or what would be the best method? Thanks!

[CharonY]

You are right, oxygen influx will severely alter the growth kinetics of your bacterium. Even if you could reach anoxic conditions faster, the influx of oxygen will either damage your cells (in case of obligates) or allow additional respiration (e.g. facultatives). An anaerobic chamber could work, and chances are that the CO2 will not make a lot of difference, unless CO2 fixation is expected.

 

What I would recommend is to use an sealed bottle for cultivation (those with large rubber stoppers) and use an gas-tight syringe that is flushed with nitrogen to draw samples for measurement. That would minimize perturbation of your culture and give more reliable results.