Watchmaker mRNA library preparation
Poly(A)-enrichment mRNA sequencing specifically isolates polyadenylated transcripts from total RNA to enable focused, strand-specific profiling of protein-coding genes, isoforms, and structural variations.
Sequencing polyadenylated messenger RNAs from total RNA is the gold standard for profiling differential gene expression, with a primary focus on protein-coding transcripts.
The Watchmaker mRNA kit is suitable wherever poly-A selected, strand-specific mRNA-seq is the right readout:
- Gene expression: Genome-wide, strand-aware quantification for differential expression studies
- Isoforms, splicing & fusions: Splice-variant identification and gene-fusion detection from full transcript coverage
- SNV detection: Single-nucleotide variant calling directly from expressed transcripts
- Novel transcript discovery: Unbiased capture of full-length poly-A transcripts
Sample requirements
- Sample type: total RNA
- Amount of material needed: 10-100 ng, method is currently validated for 100 ng input but also offered as a low input version from 10 ng
- Concentration (standard version): 2-50 ng/μL
- Concentration (low input version): 0.2-5 ng/μL
- Volume: >15 μL (max 100 µL per well). Approx 2 μL of the sample will be used for our initial quality checks, so please account for this when sending us samples
- RIN-value: >8 (RIN values may not be directly applicable for non-mammalian samples, contact us to discuss this)
- Sample extraction method: We recommend using a column-based kit without phenol. The samples should be re-suspended in nuclease-free water.
How to evaluate the sample quality
We check your samples upon arrival, however we still require our users to do their own QC steps before sending samples. To accomplish this we recommend using the following methods.
Concentration
Fluorometric measurements (Qubit, Quant-it)
Do not use absorbance measurements (Nanodrop, spectrophotometer)
RNA quality
Capillary electrophoresis (Fragment Analyzer, Bioanalyzer, TapeStation). Do not use gel electrophoresis.
If you are not able to carry out these steps, or your samples are below the required thresholds, please get in touch.
What we do with your samples
Once your samples arrive at NGI, we start by performing a reception control step in which we make sure the sample meets our requirements.
If the samples fail this quality control step, we will contact you to discuss possible options. Should you choose to proceed with samples not fulfilling the criteria it will be at your own risk, NGI will only make one attempt at library preparation and if the prep does not work you will have to pay for the library preparation anyway.
If the samples pass reception control, we will inform you and the samples will be queued for library preparation.
Library preparation
Polyadenylated messenger RNA is captured from the total RNA sample using magnetic beads. After this selection the mRNA is fragmented and cDNA is synthesised. Illumina adapters with sample specific barcode sequences are ligated and the library is amplified using PCR.
Library QC and sequencing
In this step, we evaluate the yield obtained and we will also determine the size distribution of the libraries generated. We will inform you of the QC status of each sample. Once the libraries have passed this QC step, they are normalised, pooled and queued for sequencing.
The sequencing will be carried out following the setup stated in the agreement.
Expected results
The sequencing results from each library prep will depend on the characteristics of the sample. For example, if your samples are degraded (low RIN scores) it may be better to choose a library preparation using ribosomal depletion instead, to avoid a 3′ bias of the sequencing reads.
Price examples
Library preparation is ca 1100 SEK per sample. Also see price examples page.
Last Updated: 2nd September 2026