Wednesday, June 17, 2009

I have been scratching my head at RR's comment to my last post. Why did they take forever to grow? How can I tell if there were only a few live cells, or if the strains just grow really slowly, or even both?!

I had transformed RecJ- and ExoI- MIV-competent cells with dprA- DNA,
then chosen a colony from these transformation plates and restreaked them on kan-tet plates,
then I chose colonies from the restreaks and grew them up in sBHI+KT overnight. They were cloudy in the morning, indicating they had grown.
That's where my cells came from.

I plotted a graph on Excel to help wrap my head around the information:
Sorry about the points not connecting... I'll figure that out.

Anyways, the cultures did reach their exponential phase between hours 5 and 6, but didn't feel inspired before that time. If they were growing slowly, wouldn't you expect a uniform slow growth for all hours after the start? So the growth curve would be more like a line than an exponential curve. But since the cultures did start growing rapidly in a short amount of time between hours 5 and 6, I think I may have started with a small number of viable cells. It's difficult to tell because optical density is a measure of all cells, alive or dead.

I'll be doing growth curve experiments tomorrow and Friday. Tomorrow I'll be monitoring the optical density of KW20, dprA-, RecJ-, and RecJdprA-- strains and Friday shall be KW20, dprA-, ExoI-, and ExoIdprA-- strains. I'm looking forward to learning more about the ExoI- growth curve, since I've noticed that it's always a log behind KW20 or RecJ-, for some reason.

1 comment:

Rosie Redfield said...

Try using a log scale for your OD axis. When the cells are growing exponentially the points should fall on a straight line. If ALL the points fall on a straight line, they were always growing exponentially, and growth initially appeared slow only because there were initially not very many cells growing.