help with transformation efficiency calculation! - (Apr/21/2010 )

Hi
im having trouble calculating the transformation efficiency for the competent cells. this is what i was given:

5 �l pUC18 (0.1 ng/ �l)
50 �l E. coli competent cells
Left on ice 30 minutes
Heat shocked at 42C for 45 seconds
Added 500 �l NZY broth
Place in 15 ml culture tube in shaking incubator 37C for 1 hr
Then spread 10 �l, 50 �l and 100 �l on different LB amp plates.

10 �l gave 495 colonies.

would really appreciate it if someone could help me out, the answer should be in cfu/ug units. im mainly uncertain on the amount of DNA that was in the plate.

thanks!
-sc7-

If you have 5 ul @ 0.1 ng/ ul, then you are starting with 0.5 ng of pUC18.
Once you add 500 ul NZY broth, your total volume is 5 ul + 50 ul + 500 ul = 555 ul.
And the 555 ul contains 0.5 ng of pUC18, so the concentration is 0.5 ng/555 ul = 0.0009 ng/ul.
If you spread 10 ul of 0.0009 ng/ul pUC18-containing solution on the plate, you are spreading 0.009 ng of pUC18. Converting to ug, this is 0.000009 ug.

If this 10 ul gave 495 colonies, you get 495 cfu / 0.000009 ug = XXXXX

(I left off the answer because I want to be sure you understand the reasoning - not just copy the answer - although I am sure that you would not do that ).

Hope this helps.

Mike
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im having trouble calculating the transformation efficiency for the competent cells. this is what i was given:

5 �l pUC18 (0.1 ng/ �l)
50 �l E. coli competent cells
Left on ice 30 minutes
Heat shocked at 42C for 45 seconds
Added 500 �l NZY broth
Place in 15 ml culture tube in shaking incubator 37C for 1 hr
Then spread 10 �l, 50 �l and 100 �l on different LB amp plates.

10 �l gave 495 colonies.

would really appreciate it if someone could help me out, the answer should be in cfu/ug units. im mainly uncertain on the amount of DNA that was in the plate.

thanks!
-sc7-