The use of real-time PCR analysis in a gene expression study of Alzheimer's disease post-mortem brains.
The measurement of gene expressions in brains with neurodegenerative diseases is a major area of brain research. The objective of our research was to determine whether quantitative real-time PCR could measure messenger RNA (mRNA) expression in brains with post-mortem intervals beyond 12h. In the present paper, we examined the quality of RNA from brain specimens of both Alzheimer's disease (AD) patients (n = 13) and non-demented normal control subjects (n = 6). To determine a unregulated endogenous reference gene in AD, we measured mRNA expressions of the commonly used reference genes beta-actin, 18S rRNA, and GAPDH. In addition, we determined whether post-mortem interval, brain weight, or age at death influences mRNA expression. Our real-time PCR analysis results indicate that mRNA expression can be detected in all brain specimens for beta-actin, 18S rRNA, GAPDH, and also synaptophysin, a known marker for AD. Further, using real-time PCR analysis, we found that beta-actin and 18S rRNA are differentially expressed in the brain specimens of both AD and control subjects, while GAPDH is similarly expressed in AD and control brain specimens. These findings suggest that GAPDH can be used as a endogenous reference gene in the study of AD brains. A comparative gene expression analysis also suggests that synaptophysin is down-regulated in AD brain specimens compared to control brain specimens.
Gutala
Ramana V
RV
Neurogenetics Laboratory, Neurological Sciences Institute, Oregon Health & Science University, Beaverton, OR 97006, USA.
Reddy
P Hemachandra
PH
D000199:Actins / Q000235:genetics / Q000378:metabolism; D000368:Aged; D000369:Aged, 80 and over; D000544:Alzheimer Disease / Q000235:genetics / Q000378:metabolism*; D052160:Benzothiazoles; D015152:Blotting, Northern / Q000379:methods; D001923:Brain Chemistry; D016022:Case-Control Studies; D018076:DNA, Complementary / Q000737:chemistry; D003643:Death; D003959:Diamines; D005260:Female; D015870:Gene Expression; D005786:Gene Expression Regulation; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D009929:Organ Size; D009930:Organic Chemicals / Q000378:metabolism; D011180:Postmortem Changes; D011804:Quinolines; D012333:RNA, Messenger / Q000032:analysis; D012337:RNA, Ribosomal, 18S / Q000235:genetics / Q000378:metabolism; D012044:Regression Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D016708:Synaptophysin / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D000199:Actins; D052160:Benzothiazoles; D018076:DNA, Complementary; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; D012333:RNA, Messenger; D012337:RNA, Ribosomal, 18S; D016708:Synaptophysin; C098022:SYBR Green I; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases