Growth Curve and OD - (Aug/06/2007 )
I been looking at this forum for a while, but I finally decided to sign up.
I've searched the forum and scoured the web and I cannot find an exact answer to my question.
I am growing up my bacteria (unknown marine strain) and I am having trouble getting a satisfying growth curve from OD.
So my question is how do I obtain a growth curve and more importantly a doubling time from a OD vs time graph? And if some kind of calibration is needed, what is that process?
My bacteria seems to be following its doubling time (OD doubles ~2hrs) up to an OD of 1.0 but then the OD slows down. I am not sure if that has to do with the log relation of OD or it just exits log phase that quickly. I have seen it go up to ODs of 6 so I assumed the log phase would last longer than 1.0 (maybe not tho)
can anyone please help??
really confused on the subject... thanks
can i just take the slope of my log phase of time vs OD and thats my generation time and
ln(2) / generation time = doubling time??
ok well if i get clear your subject, you need to seed 2 cultures.
Pick the OD every quarter of hour. Then you'll be able to draw your initial growth curve. you need to find where the culture is in exponential phase (linear doubling time, means your OD=f(t) curve is at this point linear too. It's here where you'll be able to truly know the doubling time.
At = Ainitial x exponential [-lambda.t]
t is in minutes, your lambda is in minute.
lambda = ln2 / T, where T is your doubling time
In grad school I was working with yeast - so I don't know if this suggestion would work - but when we wanted to do growth curves we would streak for single colonies on solid support media (YPAD plates, but I'm sure your media would need to be different). Pick a single and innoculate a flask with 50ml of liquid media in a 30oC shaking incubator. Every hour I would take a three tiny samples (100ul) and resuspend in 900ul of fresh media and take the OD600 in a 1 ml quartz cuvette (blanked against media, not water). The dilution factor was important becuase the OD600 of most cell cultures tops out and becomes non-linear fairly fast. Hope that all makes sense.
Alternatively, if you have a good microscope, you could use a hemocytometer (or something similar) to count cells directly. This is what I do now (I currently work in a mammalian cell culture lab).