Quantitative reverse transcriptase PCR to gauge increased protease-activated receptor 1 (PAR-1) mRNA copy numbers in the Wobbler mutant mouse.
Thrombin acts on cells through the surface protease-activated receptor 1 (PAR-1), a G-protein-coupled member of the seven-transmembrane domain superfamily. On neural cells, thrombin has deleterious effects, killing neurons through apoptosis. Consequently, knowledge of PAR-1 expression in the nervous system may help to elucidate the role of thrombin in neurodegenerative disease. We developed a mimic construct to facilitate the highly sensitive technique of quantitative reverse transcriptase to PCR (qRT-PCR) to measure the differential expression of low copy number PAR-1 mRNA in neurodegenerative model systems. In this article, we report our results comparing homozygous wobbler (wr/wr) mice and normal littermates. By optimizing the transcription and quantitative PCR procedures to facilitate rapid copy number determination in small RNA samples, we documented a fivefold greater level of PAR-1 mRNA in the cervical spinal cord of wr/wr.
Department of Veterans Affairs Medical Center, Kansas City, MO, USA.
R M
RM
Salcedo
B W
BW
Festoff
B A
BA
Citron
D000818:Animals; D005260:Female; D008297:Male; D051379:Mice; D008817:Mice, Mutant Strains; D018716:Molecular Mimicry; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis*; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin / Q000235:genetics*; D015183:Restriction Mapping; D013116:Spinal Cord / Q000737:chemistry
D012333:RNA, Messenger; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin