I have an LB growth media plate with E.coli on it. I want to dilute the culture. I am working with 3M Coliform detection plates, and I want to get a solution of E.coli diluted enough to grow single colonies, rather than a blob or solid streak of E.coli (on the Colliform plates). Should I measure out LB agar (with E.coli growth on it) and add it to a liquid buffer like phosphate buffer solution to make a 1:10 dilution (example .5ml E.coli media : 4.5ml PBS) and blend? Or should I transfer the E.coli culture to a liquid growth medium, incubate, and then start the dilution from that? Maybe I am thinking about this all wrong, if anyone can provide some advice that would be a great help.
Sam John Rice Donato The easiest way to handle this is to swab your original plate and dip (and mix) it in PBS. Then make a serial dilution in PBS and pipet a certain volume (+spread it out) of every tube of your serial dilution onto a plate. Atleast one of your plates will have single colonies. Good luck with your experiment and make sure your materials used are sterile before use.
I added a picture so you can visually see how it works.
Sam John Rice Donato if you want to count colonies then the dilution protocol that Laurent Lanckman provided will work great. However if you only need to get a few single colonies you can just streak isolate from the original plate to get single colonies with a sterile innoculating loop.
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