Production of knockout rats using ENU mutagenesis and a yeast-based screening assay.
The rat is a widely used model in biomedical research and is often the preferred rodent model in many areas of physiological and pathobiological research. Although many genetic tools are available for the rat, methods to produce gene-disrupted knockout rats are greatly needed. In this study, we developed protocols for creating N-ethyl-N-nitrosourea (ENU)-induced germline mutations in several rat strains. F1 preweanling pups from mutagenized Sprague Dawley (SD) male rats were then screened for functional mutations in Brca1 and Brca2 using a yeast gap-repair, ADE2-reporter truncation assay. We produced knockout rats for each of these two breast cancer suppressor genes.
Zan
Yunhong
Y
Department of Oncology, McArdle Laboratory for Cancer Research, 1400 University Avenue, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Haag
Jill D
JD
Chen
Kai-Shun
KS
Shepel
Laurie A
LA
Wigington
Don
D
Wang
Yu-Rong
YR
Hu
Rong
R
Lopez-Guajardo
Christine C
CC
Brose
Heidi L
HL
Porter
Katherine I
KI
Leonard
Rachel A
RA
Hitt
Andrew A
AA
Schommer
Stacy L
SL
Elegbede
Anu F
AF
Gould
Michael N
MN
D000818:Animals; D019313:BRCA1 Protein / Q000172:deficiency / Q000235:genetics; D024682:BRCA2 Protein / Q000172:deficiency / Q000235:genetics; D002478:Cells, Cultured; D005038:Ethylnitrosourea / Q000008:administration & dosage*; D005786:Gene Expression Regulation / Q000187:drug effects*; D018390:Gene Targeting / Q000379:methods*; D008297:Male; D016297:Mutagenesis, Site-Directed; D015202:Protein Engineering / Q000379:methods; D051381:Rats; D011922:Rats, Mutant Strains / Q000235:genetics* / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015003:Yeasts / Q000187:drug effects / Q000302:isolation & purification
D019313:BRCA1 Protein; D024682:BRCA2 Protein; D011994:Recombinant Proteins; D005038:Ethylnitrosourea