ArticleBMC molecular biology2008
Quantification of mRNA in single cells and modelling of RT-qPCR induced noise.
Article in BMC molecular biology, 2008. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 53 papers.
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Who cites it
53 citing papers in PubMed.
- Modelling HIV-1 2-LTR dynamics following raltegravir intensification.Journal of the Royal Society, Interface · 2013Trial
- Microfluidic platform for nanoliter qPCR of several retinal pigment epithelial (RPE) cells.Journal of micromechanics and microengineering : structures, devices, and systems · 2026Article
- Proteomic analysis of pancreatic endocrine cells by mechanistic single-cell isolation identifies membrane pathways.Experimental & molecular medicine · 2026Article
- Article
- RNA cytometry of single-cells using semi-permeable microcapsules.Nucleic acids research · 2023Article
- Single-Cell Mass Spectrometry Enables Insight into Heterogeneity in Infectious Disease.Analytical chemistry · 2022Article
- Semi-deconvolution of bulk and single-cell RNA-seq data with application to metastatic progression in breast cancer.Bioinformatics (Oxford, England) · 2022Article
- A protocol for extracting immunolabeled murine cardiomyocytes of high-quality RNA by laser capture microdissection.STAR protocols · 2022Article
- Tutorial: Guidelines for Single-Cell RT-qPCR.Cells · 2021Review
- Shedding light: The importance of reverse transcription efficiency standards in data interpretation.Biomolecular detection and quantification · 2019Article
- Targeted transcript quantification in single disseminated cancer cells after whole transcriptome amplification.PloS one · 2019Article
- A Combinatorial Single-cell Approach to Characterize the Molecular and Immunophenotypic Heterogeneity of Human Stem and Progenitor Populations.Journal of visualized experiments : JoVE · 2018Article
- Identification of inhibitors regulating cell proliferation and FUS-DDIT3 expression in myxoid liposarcoma using combined DNA, mRNA, and protein analyses.Laboratory investigation; a journal of technical methods and pathology · 2018Article
- Global and targeted approaches to single-cell transcriptome characterization.Briefings in functional genomics · 2018Article
- Expanded Genomic Profiling of Circulating Tumor Cells in Metastatic Breast Cancer Patients to Assess Biomarker Status and Biology Over Time (CALGB 40502 and CALGB 40503, Alliance).Clinical cancer research : an official journal of the American Association for Cancer Research · 2018Article
- Single-cell RNA sequencing to explore immune cell heterogeneity.Nature reviews. Immunology · 2018Review
- Triage of high-risk HPV-positive women in population-based screening by miRNA expression analysis in cervical scrapes; a feasibility study.Clinical epigenetics · 2018Article
- Highly multiplexed single-cell quantitative PCR.PloS one · 2018Article
- Single Cell Analysis Linking Ribosomal (r)DNA and rRNA Copy Numbers to Cell Size and Growth Rate Provides Insights into Molecular Protistan Ecology.The Journal of eukaryotic microbiology · 2017Article
- The correlation between expression profiles measured in single cells and in traditional bulk samples.Scientific reports · 2016Article
Corrections and comments
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Authors and funding
4 authors.
Funding
Abstract
backgroundGene expression has a strong stochastic element resulting in highly variable mRNA levels between individual cells, even in a seemingly homogeneous cell population. Access to fundamental information about cellular mechanisms, such as correlated gene expression, motivates measurements of multiple genes in individual cells. Quantitative reverse transcription PCR (RT-qPCR) is the most accessible method which provides sufficiently accurate measurements of mRNA in single cells.
resultsLow concentration of guanidine thiocyanate was used to fully lyse single pancreatic beta-cells followed by RT-qPCR without the need for purification. The accuracy of the measurements was determined by a quantitative noise-model of the reverse transcription and PCR. The noise is insignificant for initial copy numbers >100 while at lower copy numbers the noise intrinsic of the PCR increases sharply, eventually obscuring quantitative measurements. Importantly, the model allows us to determine the RT efficiency without using artificial RNA as a standard. The experimental setup was applied on single endocrine cells, where the technical and biological noise levels were determined.
conclusionNoise in single-cell RT-qPCR is insignificant compared to biological cell-to-cell variation in mRNA levels for medium and high abundance transcripts. To minimize the technical noise in single-cell RT-qPCR, the mRNA should be analyzed with a single RT reaction, and a single qPCR reaction per gene.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.