random Mutagenesis

Paul N Hengen pnh at fcsparc6.ncifcrf.gov
Tue Jul 25 12:51:26 EST 1995


In article <3um9ef$6on at bubba.NMSU.Edu> dkim at nmsu.edu (Daniel Kim) writes:

: I need to do random mutagenesis on a 3kb fragment.
: Is there any efficient way of doing the mutation,
: say by PCR?

| There is a method called DNA shuffling mutagenesis by a guy named Willem 
| Stemmer, published in SCIENCE and/or PNAS in late 1994/early 1995.  He 
| takes a PCR fragment, DNAse I digests it and purifies out 50 - 200 bp 
| fragments, then run PCR cycles with NO primers to let the fragments 
| anneal and primer off each other, introducing recombination errors as 
| well as PCR misincorporation errors.

@article{Stemmer1994a,
author = "W. P. Stemmer",
title = "Rapid evolution of a protein {{\em in vitro}}
by {DNA} shuffling",
journal = "Nature",
volume = "370",
pages = "389-391",
comment = "DNA shuffling mutagenesis",
year = "1994"}

@article{Stemmer1994b,
author = "W. P. Stemmer",
title = "{DNA} shuffling by random fragmentation and reassembly:
{{\em in vitro}} recombination for molecular evolution",
journal = "Proc. Natl. Acad. Sci. USA",
volume = "91",
pages = "10747-10751",
comment = "DNA shuffling mutagenesis",
year = "1994"}

I would be interested to hear the experiences of anyone who
has tried this procedure.

*******************************************************************************
* Paul N. Hengen, Ph.D.                           /--------------------------/*
* National Cancer Institute                       |Internet: pnh at ncifcrf.gov |*
* Laboratory of Mathematical Biology              |   Phone: (301) 846-5581  |*
* Frederick Cancer Research and Development Center|     FAX: (301) 846-5598  |*
* Frederick, Maryland 21702-1201 USA              /--------------------------/*
* - - - - - Methods FAQ list -> http://www-lmmb.ncifcrf.gov/~pnh/ - - - - - - *
* - - -  Anonymous FTP from ftp.ncifcrf.gov as file pub/methods/FAQlist - - - *
*******************************************************************************



More information about the Methods mailing list