one-step gene disruptions

Preston Garrison garrisonp at THORIN.UTHSCSA.EDU
Tue Apr 19 15:01:01 EST 1994


Dave Eide wrote:
>Buddies,
>
>We would like to do some one-step gene disruptions using flanking regions 
>smaller than the recommended minimum.  Rothstein (Meth. Enzymol. vol. 194) 
>suggests using flanking regions of 100 bp or more on each side of the
selectable
>marker and the method we want to try gives us only about 50 bp on each side.  
>Does anyone have information regarding homologous recombination of fragments of

>this size?  Any input will be greatly appreciated.
>
>Dave Eide 
>
It is possible to get targeting with sequences that short. See Baudin et al.
Nucl. Acid Res. 21: 3329 1993. Also, Fred Sherman had several papers a few
years ago describing transformation with oligos that restored function to 
mutant cytochrome c genes. The efficiency of targeting to the intended site is 
lower than you get with larger homologous regions.

Preston Garrison
Biochem. Dept.
UTHSCSA
San Antonio, Tx 78284-7760
USA
garrisonp at uthscsa.edu



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