The measurement of adenosine and estrogen receptor expression in rat brains following ovariectomy using quantitative PCR analysis
Author(s)
Rose'Meyer, RB
Mellick, AS
Garnham, BG
Harrison, GJ
Massa, HM
Griffiths, LR
Griffith University Author(s)
Year published
2003
Metadata
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In our laboratory we have developed a quantitative-polymerase chain reaction (Q-PCR) strategy to examine the differential expression of adenosine receptor (ADOR), A1, A2A, A2B and A3, and estrogen receptors (ER) a and ߮ Brain and uterine mRNA were first used to optimise specific amplification conditions prior to SYBR Green I real time analysis of receptor subtype expression. SYBR Green I provided a convenient and sensitive means of examining specific PCR amplification product in real time, and allowed the generation of standard curves from which relative receptor abundance could be determined. Real time Q-PCR analysis was ...
View more >In our laboratory we have developed a quantitative-polymerase chain reaction (Q-PCR) strategy to examine the differential expression of adenosine receptor (ADOR), A1, A2A, A2B and A3, and estrogen receptors (ER) a and ߮ Brain and uterine mRNA were first used to optimise specific amplification conditions prior to SYBR Green I real time analysis of receptor subtype expression. SYBR Green I provided a convenient and sensitive means of examining specific PCR amplification product in real time, and allowed the generation of standard curves from which relative receptor abundance could be determined. Real time Q-PCR analysis was then performed, to examine changes in receptor expression levels in brains of adult female Wistar rats 3-month post ovariectomy. Comparison with sham-operated age-matched control rats demonstrated both comparative and absolute-copy number changes in receptor levels. Evaluation of both analytical methods investigated 18S rRNA as an internal reference for comparative gene expression analysis in the brain. The results of this study revealed preferential repression of ADORA2A (>4-fold down) and consistent (>2-fold) down-regulation of ADORA1, ADORA3, and ER-߬ following ovariectomy. No change was found in ADORA2B or ER-a. Analysis of absolute copy number in this study revealed a correlation between receptor expression in response to ovariectomy, and relative receptor subtype abundance in the brain.
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View more >In our laboratory we have developed a quantitative-polymerase chain reaction (Q-PCR) strategy to examine the differential expression of adenosine receptor (ADOR), A1, A2A, A2B and A3, and estrogen receptors (ER) a and ߮ Brain and uterine mRNA were first used to optimise specific amplification conditions prior to SYBR Green I real time analysis of receptor subtype expression. SYBR Green I provided a convenient and sensitive means of examining specific PCR amplification product in real time, and allowed the generation of standard curves from which relative receptor abundance could be determined. Real time Q-PCR analysis was then performed, to examine changes in receptor expression levels in brains of adult female Wistar rats 3-month post ovariectomy. Comparison with sham-operated age-matched control rats demonstrated both comparative and absolute-copy number changes in receptor levels. Evaluation of both analytical methods investigated 18S rRNA as an internal reference for comparative gene expression analysis in the brain. The results of this study revealed preferential repression of ADORA2A (>4-fold down) and consistent (>2-fold) down-regulation of ADORA1, ADORA3, and ER-߬ following ovariectomy. No change was found in ADORA2B or ER-a. Analysis of absolute copy number in this study revealed a correlation between receptor expression in response to ovariectomy, and relative receptor subtype abundance in the brain.
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Journal Title
Brain Research Protocols
Volume
11
Publisher URI
Copyright Statement
© 2003 Elsevier : Reproduced in accordance with the copyright policy of the publisher : This journal is available online - use hypertext links
Subject
Neurosciences
Psychology
Cognitive Sciences