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Is it possible to compare RPKM values across studies? This question is something I have been thinking about during my own analyses. I tend to think that under the conditions of standard poly-A enrichment and approximately equal coverage during sequencing runs, then RPKM values for genes between separate studies can be compared. However, if study X used a transcript enrichment setup in which a select number of mRNAs from a subset of genes were being selected for sequencing, can the RPKM values obtained from this study be reliably compared to study Y in which poly-A enrichment was used?
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Part II for those who are feeling ambitious: If I were to selectively enrich for a set of transcripts (A,B,C), sequence these, and obtain high RPKM measurements and attempt to compare these to a poly-A enriched sample set in which I have a fair expression measurement of most of the transcriptome (although in this case transcripts A, B, and C were unaccounted for due to low abundance) is it possible to extrapolate the normal physiological mRNA abundance of these transcripts from the enriched data series?
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Data variability is still an issue in sequencing technologies despite the advances in reproducibility and accuracy of these platforms. But the problem does not originate in the sequencing itself, but in the previous steps, before the sample reaches the sequencer.
The first step is collection, followed by preservation and sample preparation for analysis. Most scientists overlook those steps, but not being careful might just be skewing the experiment’s results.
...06-02-2026, 10:05 AM -
With the launch of new single-cell sequencing platforms in 2026, the field stands at an exciting inflection point. This article surveys the most impactful advances in the field and discusses how they’re reshaping research in cancer, immunology, and beyond.
Introduction
Single-cell sequencing technologies have undergone remarkable advances over the past decade, transitioning from low-throughput experimental approaches to highly scalable platforms capable of...05-22-2026, 06:42 AM
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