cDNA library amplification

Warren Gallin wgallin at gpu.srv.ualberta.ca
Mon Jan 14 11:20:26 EST 2002


Kevin,
	If the library is unamplified, then the titer times the volume must
equal the complexity.
	Since it does not, you are either looking at an amplified library, or
some your numbers are way off.

Warren Gallin

Kevin O'Connell wrote:
> 
> Hi All,
> 
> I am hoping that someone will be able to help me with what should be a
> simple procedure.  I have a 1.5 ml aliquot of an unamplified two-hybrid cDNA
> phage library whose titer is purported to be 10e10 per ml.  However I have
> titered it myself and found it to be 6x10e7.  The person who constructed it
> claims that it has 10e7 independent clones.  I wish to amplify the library
> and it seems to me that I would have to use the entire 1.5 ml aliquot or
> else the amplified library would have far fewer independent clones than the
> original.  The questions are: Can I or should I use the entire 1.5 ml and if
> I do, what volume of a log stage bacterial culture would I need to use.
> Also what is the maximal number of PFU that can be plated on a 150 mm plate.
> That is how many plates would I need.  One protocol call for 10e5 per 150 mm
> plate but if I used the entire aliquot I would have to use over 100 plates
> and this is obviously way too many.  If anybody has some suggestions, I
> would greatly appreciate it.
> 
> Kevin




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