2000 cfu/plate - how to prepare LB-Plates with 2000 cfu? (Mar/10/2008 )

Hello everybody,

i like to prepare some LB-Plates on which i should have an amount of 2000 single colonies. How can i do that? Measure OD600 and calculate (P. syringae)?
Or some kind of dilutions?
Help me! Please!

thx for help,
thisastar

The optimal number of colonies per plate is between 30 and 300. Below 30 colonies, the standard deviation is too high and above 300 it is difficult to pick single colonies. Dilute your cells to get a number between 30 and 300 and prepare enough plates to get the total of 2000 colonies you need.

I will need the big plates (r=72.5 mm). And i like to do some replica plates, therefore i need a lot of colonies. with 2000 colonies i would have all 8mm one (if i calculated correctly ). the replica plating i'll do with some kind of stamp, means i dont pick single colonies.
but back to the orignial question: how can i know how much of a culture i need to plate out to get 2000 (or any number) on my plate? because when i plate them out for a kind of a dilution row, they need to grow. and in this time my liquid culture grows too. so then the number isnt anymore correct. either i'm blind for a simple solution of that much stupid...

grze

Measure the OD, do 10-fold series dilutions, plate it out and count.

bascially do a trial run.

Innoculate 2ml of culture with a single colony or Y amount of cells, and grow for X amount of time. Do a serial dilution. Plate on normal plates. Count cells. THen you can work out how much of your culture that you need to plate.

One you got the values, then do the big experiment.

thx to all of u,

i'll try to dilute after OD600 measurement, i calculate then with 10^8 cells/ml on an optical density of 0.5

greez