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Published on: February 8, 2013
RNA degradation in human breast tissue after surgical removal: a time-course study
Yoko Ohashi1, Kim E Creek, Lucia Pirisi
1Department of Obstetrics and Gynecology, University of South Carolina School of Medicine, Columbia, SC 29203, USA.
Abstract:
There is much interest in the study of human malignancy using gene expression profiling techniques. Expression profiles obtained from microarrays utilize RNA extracted from the tissue in question. Currently, cell cultures or fresh tissue processed "quickly" are used in these studies. To our knowledge, there are no published reports of a time-course of RNA degradation in surgically removed breast tissue. Such a time-course study is critically needed. We obtained normal breast tissue from breast reduction surgery. Portions of breast tissue kept at room temperature were sampled and placed into RNAlater to preserve RNA at different time-points from 10 min to 3 h after the surgical removal. We evaluated total RNA integrity from each specimen using agarose gel electrophoresis and real-time quantitative RT-PCR analysis of four genes. Electrophoresis showed good-quality, intact RNA at all time points up to 3 h. Quantitative RT-PCR showed no difference in amplified products among all samples. Our study showed that there was no loss of RNA integrity in normal breast tissue for up to 3 h after surgical removal.
Insights
Normal breast tissue retains high RNA integrity for up to 3 hours post-surgery at room temperature. This finding is crucial for gene expression profiling in breast cancer research.
Area of Science:
- Molecular Biology
- Oncology
- Biochemistry
Background:
- Gene expression profiling is vital for studying human malignancy.
- Current methods for RNA extraction often rely on cell cultures or immediate processing of fresh tissue.
- Limited data exists on RNA stability in surgically removed breast tissue over time.
Purpose of the Study:
- To investigate the time-course of RNA degradation in normal breast tissue after surgical removal.
- To determine the optimal time window for RNA preservation without specialized handling.
- To provide essential data for optimizing sample collection protocols in breast cancer research.
Main Methods:
- Normal breast tissue samples were collected from reduction surgeries.
- Tissue portions were stored at room temperature and sampled at intervals from 10 minutes to 3 hours.
- RNA preservation was achieved using RNAlater solution.
- RNA integrity was assessed via agarose gel electrophoresis.
- Gene expression levels were quantified using real-time quantitative RT-PCR for four genes.
Main Results:
- Agarose gel electrophoresis confirmed good-quality, intact RNA across all time points up to 3 hours.
- Real-time quantitative RT-PCR analysis showed no significant differences in amplified products between samples.
- RNA integrity remained stable in normal breast tissue for up to 3 hours post-excision at room temperature.
Conclusions:
- Normal breast tissue maintains RNA integrity for at least 3 hours after surgical removal when stored at room temperature.
- These findings support the feasibility of delayed RNA processing for gene expression studies using breast tissue.
- The study provides a critical time-course data for establishing reliable sample handling protocols in breast cancer research.
