Genotyping microsatellite polymorphisms by agarose gel electrophoresis with ethidium bromide staining: application to quantitative trait loci analysis of seizure susceptibility in mice.
Agarose gel electrophoresis with ethidium bromide staining (AGE/EBS) is an efficient and reliable method for analyzing microsatellite polymorphisms. We report the use of AGE/EBS for analyzing DNA microsatellite polymorphisms in a preliminary quantitative trait loci (QTL) study of seizure susceptibility in which a candidate gene strategy was used to direct initial mapping efforts. F2 intercross progeny, derived from seizure-sensitive DBA/2J (D2) and seizure-resistant C57BL/6J (B6) inbred strains of mice, were tested for their sensitivity to the seizure-inducing effect of pentylenetetrazol (PTZ), a gamma-aminobutyric acid (GABA) receptor antagonist. A semi-automated method is described, in which DNA microsatellites were amplified by polymerase chain reaction (PCR) to yield products of 100-200 base pair (bp) in length. Alleles were separated on 3-6% MetaPhor agarose gels, stained with ethidium bromide, and visualized by ultraviolet (UV) illumination. Univariate analysis of genotype and phenotype data provides evidence for a seizure-related QTL on chromosome 5, near genes coding for the GABAA receptor subunits alpha 5 and gamma 3. Interestingly, this suggestive QTL derives from the more resistant B6 strain, but it nonetheless provides impetus for the characterization of possible strain differences in these two candidate genes. Overall, these results demonstrate that AGE/EBS can be useful for rapid screening of genomic regions of special interest in QTL mapping studies.
Ferraro
T N
TN
Department of Psychiatry, Center for Neurobiology and Behavior, University of Pennsylvania, Philadelphia 19104, USA. tnf@mail.med.upenn.edu
Schill
J F
JF
Ballas
C
C
Mulholland
N
N
Golden
G T
GT
Smith
G G
GG
Buono
R J
RJ
Berrettini
W H
WH
D000818:Animals; D002874:Chromosome Mapping; D003292:Convulsants / Q000633:toxicity; D003433:Crosses, Genetic; D004247:DNA / Q000032:analysis / Q000235:genetics; D004351:Drug Resistance; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004996:Ethidium; D005260:Female; D005456:Fluorescent Dyes; D058787:GABA-A Receptor Antagonists; D020022:Genetic Predisposition to Disease; D005838:Genotype; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL / Q000235:genetics*; D008811:Mice, Inbred DBA / Q000235:genetics*; D018895:Microsatellite Repeats; D010433:Pentylenetetrazole / Q000633:toxicity; D019655:Quantitative Trait, Heritable; D011963:Receptors, GABA-A / Q000235:genetics*; D012640:Seizures / Q000139:chemically induced / Q000235:genetics*; D013194:Staining and Labeling
D003292:Convulsants; D005456:Fluorescent Dyes; D058787:GABA-A Receptor Antagonists; D011963:Receptors, GABA-A; D004247:DNA; D004996:Ethidium; D010433:Pentylenetetrazole