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Quantitation of mRNA levels of steroid 5α-reductase isozymes in the rat brain by “one-step” RT-PCR and capillary electrophoresis
| Content Provider | Semantic Scholar |
|---|---|
| Author | Torres, Jesús M. Sánchez, Pilar Domínguez Ortega, Esperanza |
| Copyright Year | 2004 |
| Abstract | The enzyme 5α-reductase (5α-R) is present in many mammalian tissues, including the brain. The physiological importance of 5α-R in the brain derives from its capability to convert testosterone (T) to a more potent androgen, dihydrotestosterone (DHT), and to convert progesterone to its 5α-reduced derivative, precursors of allopregnanolone, potent allosteric modulator of the γ-aminobutyric acid receptor (GABAA-R). 5α-R occurs as two isoforms, 5α-R type 1 (5α-R1) and 5α-R type 2 (5α-R2). We present an accurate, rapid, and modestly labor-intensive method to precisely quantitate 5α-R mRNA species in the cerebral cortex of the rat. This approach combines the high specificity of “one-step” reverse transcription-polymerase chain reaction (RT-PCR) with the sensitivity of laser-induced fluorescence capillary electrophoresis (LIF-CE). Both cDNA synthesis and PCR amplification are performed with the same enzyme and site-specific primers, improving the efficiency of cDNA synthesis. The specific target mRNA and a mimic DNA fragment, used as a competitive internal standard, were co-amplified in a single reaction in which the same primers are used. The method presented in this paper enables a more efficient quantitative determination of 5α-R mRNA isozymes, and may lead to a better understanding of the role of 5α-R isozymes in the physiology of the central nervous system. |
| Starting Page | 211 |
| Ending Page | 216 |
| Page Count | 6 |
| File Format | PDF HTM / HTML |
| DOI | 10.1016/j.jneumeth.2003.12.014 |
| Volume Number | 135 |
| Alternate Webpage(s) | https://api.elsevier.com/content/article/pii/S0165027003004357 |
| Alternate Webpage(s) | https://www.sciencedirect.com/science/article/pii/S0165027003004357?dgcid=api_sd_search-api-endpoint |
| Alternate Webpage(s) | https://doi.org/10.1016/j.jneumeth.2003.12.014 |
| Journal | Journal of Neuroscience Methods |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
