Background: Although TRIzol is widely used for preservation and isolation of RNA, there is suspicion that prolonged sample storage in TRIzol may affect array-based gene expression profiling (GEP) through premature termination during reverse transcription.
Methods: GEP on Illumina arrays compared paired aliquots (cryopreserved or stored in TRIzol) of primary samples of multiple myeloma (MM) and acute myeloid leukemia (AML). Data were analyzed at the "probe level" (a single consensus value) or "bead level" (multiple measurements provided by individual beads).
Results: TRIzol storage does not affect standard probe-level comparisons between sample groups: different preservation methods did not generate differentially expressed probes (DEP) within MM or AML sample groups, or substantially affect the many DEPs distinguishing between these groups. Differences were found by gene set enrichment analysis, but these were dismissible because of instability with permutation of sample labels, unbalanced restriction to TRIzol aliquots, inconsistency between MM and AML groups, and lack of biological plausibility. Bead-level comparisons found many DEPs within sample pairs, but most (73%) were <2-fold changed. There was no consistent evidence that TRIzol causes premature reverse transcription termination. Instead, a subset of DEPs were systematically due to increased signals in TRIzol-preserved samples from probes near the 5' end of transcripts, suggesting better mRNA preservation with TRIzol.
Conclusions: TRIzol preserves RNA quality well, without a deleterious effect on GEP. Samples stored frozen with and without TRIzol may be compared by GEP with only minor concern for systematic artifacts.
Impact: The standard practice of prolonged sample storage in TRIzol is suitable for GEP.
Download full-text PDF |
Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2966770 | PMC |
| http://dx.doi.org/10.1158/1055-9965.EPI-10-0565 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
Factors influencing RNA yield from placenta tissue.
Placenta
September 2023
Flinders Health and Medical Research Institute, Flinders University, Adelaide, South Australia, Australia. Electronic address:
High yield and integrity of placental RNA are crucial for placental transcriptomics studies. We assessed the effects of time to placental collection post-delivery; tissue storage, amount and method used for extraction; mode of delivery; and tissue type on total RNA yield. The optimal protocol for RNA extraction from placental tissue includes cryofreezing of the sample upon collection and RNA extraction from 50 mg of tissue using TRIzol reagent.
View Article and Find Full Text PDFComparison of Four Methods of RNA Extraction and cDNA Synthesis from The Venom of Peruvian Snakes of the Genus of Clinical Importance.
Int J Mol Sci
July 2023
Laboratorio de Biología Molecular, Facultad de Ciencias Biológicas, Universidad Nacional Mayor de San Marcos, Av. Venezuela Cdra 34 S/N, Ciudad Universitaria, Lima Cercado, Lima 15081, Peru.
RNA purification and cDNA synthesis represents the starting point for molecular analyses of snake venom proteins-enzymes. Usually, the sacrifice of snakes is necessary for venom gland extraction to identify protein-coding transcripts; however, the venom can be used as a source of transcripts. Although there are methods for obtaining RNA from venom, no comparative analysis has been conducted in the genus.
View Article and Find Full Text PDFStability investigation of air-dried olive ribo nucleic acids for metavirome studies.
Plant Methods
February 2022
Temperate Fruit Research Center, Horticultural Research Institute, Agricultural Research, Education and Extension Organization (AREEO), P. O. Box: 31585-4119, Karaj, Iran.
Background: The application of ribo nucleic acids for molecular studies requires high integrity and quality of extracted total RNA samples. In addition, the need to transfer RNA samples at room temperature without special treatments such as ice and liquid nitrogen storage according to international transport laws highlights the importance of low cost alternative methods such as RNA air-drying, lyophilisation and transportable agents. In this study, the quality and quantity of air-dried RNA samples from leaf, petiole and bark tissues of different olive genotypes using several RNA extraction methods were compared with lyophilized ground leaves and RNAlater-stored tissue samples before precipitation.
View Article and Find Full Text PDFComparison of methods for pre-processing, exosome isolation, and RNA extraction in unpasteurized bovine and human milk.
PLoS One
November 2021
Center for Environmental Epigenetics and Development, Department of Biological Sciences, University of Toronto Scarborough, Toronto, Ontario, Canada.
Milk is a highly complex, heterogeneous biological fluid that contains non-nutritive, bioactive extracellular vesicles called exosomes. Characterization of milk-derived exosomes (MDEs) is challenging due to the lack of standardized methods that are currently being used for milk pre-processing, storage, and exosome isolation. In this study, we tested: 1) three pre-processing methods to remove cream, fat, cellular debris, and casein proteins from bovine milk to determine whether pre-processing of whole milk prior to long-term storage improves MDE isolations, 2) the suitability of two standard exosome isolation methods for MDE fractionation, and 3) four extraction protocols for obtaining high quality RNA from bovine and human MDEs.
View Article and Find Full Text PDFDevelopment of an alternative saliva test for diagnosis of SARS-CoV-2 using TRIzol: Adapting to countries with lower incomes looking for a large-scale detection program.
PLoS One
August 2021
Centro de Medicina Tropical, División de Investigación, Facultad de Medicina, Universidad Nacional Autónoma de México, Mexico City, Mexico.
Article Synopsis
- Saliva testing for SARS-CoV-2 is a promising alternative to traditional nasopharyngeal swabs, allowing for self-collection and reducing the risk of exposure to the virus.
- Current saliva tests are hindered by high costs due to the need for specialized tubes, prompting the development of a method using TRIzol to stabilize RNA and inactivate the virus.
- The new TRIzol saliva testing method has been shown to be just as effective as traditional swabs, offers greater accessibility, is less expensive, and is particularly beneficial for low-income countries in managing COVID-19 cases.
Want AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!