Calculation of transformation efficiency

David L. Haviland, Ph.D. dhavilan at IMM2.IMM.UTH.TMC.EDU
Tue Feb 4 10:52:17 EST 1997


At 12:51 2/4/97 +0000, Kevin A. Mulcahy wrote:
>I have been trying to work out how to estimate transformation=20
>efficiencies but in my searching through various texts I have found 3=20
>different (and confusing!) ways which all give different answers!. Could=20
>you please tell me if the calculation written out below is correct?
>
>200=B5l of E. coli are transformed with 1ng of pBR322, made up to 1ml with=
=20
>growth medium for the recovery phase and then 10=B5l of this plated plated=
=20
>out (i.e. 1% of the total transformation mixture, therefore equivalent=20
>to 10pg DNA used) and 100 colonies obtained. Is the transformation=20
>efficiency 1 x 10(exp)7 cfu/=B5g DNA? (i.e. 100 colonies per 10pg DNA and=
=20
>therefore 10,000,000 colonies per =B5g of DNA).

A while ago I too had had this trouble too and one day I just broke down
and asked various folk in the tech-services departments of Promega and
Stratagene.  I happily obtained consistient answers and for your
application would be:

100 cfu/1ng

times 1000ul-original volume/ 10ul plated  volume  - correction to 1 ml=
 (100)

times  1000ng/ug

equals 1 x 10^7 cfu/ug  -- you're doing it right!   A respectible
effeciency...

When the companies "punch it out" with their modified Hanahan protocol
(proprietary modification, of course!!), they are transforming a range of
DNA concentrations in the nanogram range.  With thier transformation
efficency they end up with an extra 100-1000 fold increase in colonies.

If you had transformed all of 50 pg and got 100 colonies, you'd be bucking
2 X 10^8!!!

The best I've obtained with Inoue's protocol has been 1-2 X10^7 cfu/ug.
I've scratched mid 10^7 to low 10^8 with electroporation.  Standard generic
CaCl2 alone, for me, yeilds low 10^4 cfu/ug and has never been higher.

Hope this helps,
David









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