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Stabilize Difficult-to-work-with RNA Transcripts
RNA stabilization during sample collection for RNA-based gene expression studies is often critical for detecting cellular and tissue responses to a stimulus. As Rona Wang and DaiWei Wang (University of British Columbia) discovered, RNAlater® Tissue Collection: RNA Stabilization Solution provides a method for preserving RNA in tissue samples for large-scale experiments. This allowed them to examine the effects of cigarette smoke on tracheal gene expression.
Rona Wang and DaiWei Wang
Department of Pathology and Department of Science, University of British Columbia, G-223-2211 Wesbrook Mall, Vancouver, BC, V6T 2B5, Canada
Email: rdwang@interchange.ubc.ca
Rona Wang and DaiWei Wang from the laboratories of Drs. A Churg and JL Wright (University of British Columbia), work with rat models to examine lung diseases related to cigarette smoking. They routinely assay changes in expression of more than a dozen genes, including procollagen type I, connective tissue growth factor (CTGF), transforming growth factor-α (TGF-α), and platelet-derived growth factor (PDGF)-α and –β, which all require that RNA extraction and RT-PCR be performed immediately after sample collection.
During a large experiment, when samples were too numerous for immediate processing, they stored rat trachea samples in Ambion RNAlater® Tissue Collection: RNA Stabilization Solution. After 1 week of storage, RNA was extracted and tested by RT-PCR for changes in gene expression. Comparable PCR signals were obtained from cDNA generated from tissues stored in RNAlater Solution and from fresh samples that were processed immediately after harvest (Figure 1). As shown in Figure 2, RT-PCR using RNA extracted from tissue stored in RNAlater solution, demonstrated that procollagen type I gene expression could still be detected even 1 week after initial organ excision and storage.
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| Figure 1. RNAlater® Tissue Collection: RNA Stabilization Solution Stabilizes Gene Expression in Rat Explants. Rat explants were either processed immediately after harvesting (Fresh) or stored in RNAlater Tissue Collection: RNA Stabilization Solution (RNAlater-treated) for 1 week. Total RNA was extracted and reverse transcribed, and the resulting cDNA was stored at –20°C for 3 months before being assayed by PCR. The control was PCR products stored at 4°C for 3 months. |
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| Figure 2. RNAlater® Tissue Collection: RNA Stabilization Solution Stabilizes Gene Expression in Rat Explants. Rat explants were either frozen at –80°C for 1 week or stored in RNAlater Tissue Collection:RNA Stabilization Solution for 1 week. Total RNA was extracted and stored at –20°C for 1 week before being used in an RT-PCR assay to detect procollagen type I. |
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