I first retrieved the yeast from the -80 degree Celsius freezer and then place it onto YM agar plate and then stored at 30 degree Celsius. Next day I took a colony from that plate and added it to another plate and stored at 30 degree Celsius. After 24 hours I took one colony from the second plate and placed it in 100 ml of liquid medium and put it in a shaking incubator at 30 degree C at 150 rpm. After 24 hours I took the broth and centrifuged it at 4000 rpm and got rid of the supernatant and weighed the pellet. The pellet should weigh 1 g and then I added 99 ml of fresh medium to make 1% yeast from the 1% I take 1 ml and add it into a new 100 ml of fresh medium and this is my 0 hours. From there I just take samples at 3 hour intervals I monitor OD and CFU. The numbers always seem weird. Really hoping for a help. Thank you.
Hi Dan
Considering you have transfered cells from -80ºC and then to agar plates repeatedly cells should be competent to grow well.
As for the standard growth curve. Which kind of medium are you using? And what do you mean by numbers don't match up? Could you give an example?
I'd suggest the following.
For what i've seen, your yeast strain can grow in minimal medium supplemented with ammonium sulphate, glucose and maybe any amino acid for which your strain may be an auxotroph. For growth curves these are the best conditions, since it's a medium which doesn't allow for instance explosive growth and in turn lets you follow the curve in a steady way.
I'd grow the yeast in a pre-innoculum overnight. In the morning i'd measure the OD and prepare a new innoculum with a OD of 0.1 (no need to wight it in my book). Then you just need to follow the growing of the yeast. I'd take OD measurings each hour for example and see how it goes and then adapt from it.
Hope it helps
Best
Rui Filipe Ramos Santos thank you for your answer, I am using YM medium. What I meant by why my numbers don't match up is that for example the CFU/ml for 6 hours maybe higher than that of 9 hours. It follows this pattern and I have tried it many times and still not sure what the problem is.
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