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Isolation of differentially expressed sequence tags from steroid-responsive cells using mRNA differential display
1Department of Biochemistry, University of Arizona, Tucson 85724, USA.
Molecular and Cellular Endocrinology
|February 27, 1995
Summary
Researchers utilized mRNA differential display reverse transcriptase PCR (DDPCR) to discover novel steroid-regulated genes in mouse thymocytes and rat prostate cells. This method identified key genes like GIG10 and GIG18, rapidly induced by dexamethasone.
Area of Science:
- Molecular Biology
- Genomics
- Endocrinology
Background:
- Steroid receptors regulate gene networks, but specific target genes are poorly understood.
- Identifying cell-specific steroid-regulated genes is crucial for understanding hormonal effects.
Purpose of the Study:
- To identify and isolate novel steroid-regulated genes using mRNA differential display reverse transcriptase PCR (DDPCR).
- To characterize differentially expressed sequence tags (DESTs) in hormone-modulated cells.
Main Methods:
- Applied mRNA differential display reverse transcriptase PCR (DDPCR) to mouse thymocytes (WEHI 7.2) and rat ventral prostate tissue.
- Isolated and characterized differentially expressed sequence tags (DESTs) under various hormone conditions.
- Modified DDPCR protocol to reduce false-positive results.
Main Results:
- Identified several DESTs in mouse thymocytes, including GIG10 (novel) and GIG18 (mouse homologue of human EST).
- GIG10 and GIG18 were rapidly induced by dexamethasone within 2 hours.
- Isolated and characterized an androgen-regulated DEST from rat prostate corresponding to prostatic spermine binding protein mRNA.
Conclusions:
- DDPCR is an effective method for discovering cell-specific, hormone-regulated genes.
- Identified novel dexamethasone-induced genes (GIG10, GIG18) and an androgen-regulated gene in relevant tissues.
- Modified DDPCR protocol enhances reliability for gene discovery.
