Amplifying genomic libraries: How many PFU's per plate?

kim at m44.unm.edu kim at m44.unm.edu
Tue Jul 2 12:10:28 EST 1991


In article <1991Jun27.220747.16895 at ccu.umanitoba.ca>, frist at ccu.umanitoba.ca writes:
>I am preparing to screen a pea genomic library in Lambda L47.1. With an 
>average insert size of 20kb, approximately 1.2e6 clones are needed to
>have a 99% chance of finding a single copy gene.
                      (stuff omitted)
>I haven't amplified a genomic library before, but I don't recall hearing
>about this scale of operation from others who have.
                       (More omitted)

First, I am not sure that one can generate adequate representation by simply
amplifying a library to get more pfu.  I always assumed that the calculated
minimum pfu for a representative library referred to the size of the library
before amplification.  That is, a "library" that has initially only 10
recombinant plaques can be amplified to contain millions of pfu, but still not
contain any more genetic information.  Does anyone else have thoughts on this?

Second, one can plate a lambda library at very high densities for amplification
if the plate lysate is harvested before the plaques are large enough to touch
each other.  This may mean harvesting at a time before plaques are really
visible, which requires a leap of faith.

Daniel Kim                                    



More information about the Methods mailing list