The Challenges of RT

The Challenges of Reverse Transcription (RT)

For years, scientists have relied on platforms that use reverse transcriptase to convert RNA to cDNA in order to perform gene expression studies. However, the cDNA conversion and amplification steps can introduce variability into the data. Additionally, other steps in the workflow like sample prep and data analysis can introduce variability. The scientific community is now recognizing these challenges, and publications on the limitations of RT-based platforms are emerging across the qPCR, microarray and NGS spaces.


The Benefits of Counting RNA Directly

What if you could bypass a cDNA conversion step and count RNA molecules directly? The nCounter® platform offers direct RNA detection for robust, reproducible performance and unbiased transcript quantitation.

Interested in learning more? Your journey begins here


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Bustin, S. et. al., “Talking the talk but not walking the walk: RT-qPCR as a paradigm for the lack of reproducibility in molecular research.” Eur J Clin Invest 2017 Oct;47(10):756-774

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Bustin, S. et. al., “Variability of the Reverse Transcription Step: Practical Implications.” Clinical Chemistry 61:1 000-000 (2015)

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Dijkstra, J.R. et. al., “Critical appraisal of quantitative PCR results in colorectal cancer research: Can we ply on published qPCR results?” Mol Oncol 2014 Jun:8(4):813-8

Esteve-Codina, A. et. al., “A Comparison of RNA-Seq Results from Paired Formalin-Fixed Paraffin-Embedded and Fresh-Frozen Glioblastoma Tissue Samples.” PLOS. January 25, 2017

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Omolo, B. et. al., “Adaptation of a RAS pathway activation signature from FF to FFPE tissues in colorectal cancer.” BMC Medical Genomics 19 October 2016.

Raman, A.T. et. al., “Apparent bias towards long gene misregulation in MeCP2 syndromes disappears after controlling for baseline variations.” Nature Communications  13 August 2018 Article number: 3225 (2018)

Roy, S. et. al., “Standards and Guidelines for Validating Next-Generation Sequencing Bioinformatics Pipelines, A Joint Recommendation of the Association for Molecular Pathology and the College of American Pathologists.” The Journal of Molecular Diagnostics Vol. 20, No. 1, January 2018

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Veldman-Jones, M.H. et. al., “Evaluating Robustness and Sensitivity of the NanoString Technologies nCounter Platform to Enable Multiplexed Gene Expression Analysis of Clinical Samples.” Cancer Res June 11, 2015: 75(13):2587-93


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