SMARTer ThruPLEX DNA-seq

Library preparation for DNA, ideal for preparing libraries from small amounts of input material. Works well for shotgun libraries, ChIP DNA and FFPE samples, amongst others.

The ThruPLEX DNA-seq chemistry is engineered and optimized to generate DNA libraries with high molecular complexity from low-input samples.

Sample requirements for common sample types

Genomic DNA

• Sample type: DNA
• Concentration: 1-10 ng/μL
• Volume requirement NGI Stockholm: >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.
• Volume requirement NGI Uppsala: 25-50 µl.
  Please contact a coordinator if the sample requirements cannot be fulfilled
• Material needed for 1 prep: 10-15 ng

FFPE

• Sample type: DNA
• Concentration: 10-50 ng/μL
• Volume: >15 μL. Approx 2 μL of your sample will be used for our initial quality measurements.
• Material needed for 1 prep: 100 ng
• We ask for: Gel picture or FragmentAnalyzer trace

ChIP-ed DNA

• Sample type: ChIP-ed DNA
• Concentration: 0.1-10 ng/μL (concentrations can very well be much lower than 0.1 ng/μL for this type of samples and this is usually fine. Limitations in our concentration measurement assay means we can not reliably quantify samples lower than 0.1 ng/μL, and thus they will be classified as failed in our QC)
• Volume: >15 μL. Approx 2 μL of your sample will be used for our initial quality measurements.
• Material needed for 1 prep: 0.1-1 ng
• We ask for: BioAnalyzer trace

These input requirements are for illustrative purposes only. Please get in touch with one of our project coordinators to discuss your samples if in doubt.

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, the samples will be queued for library preparation.

Library preparation

Single-tube reaction that starts with low input double-stranded DNA (0.05 ng to 50 ng). The genomic DNA is fragmented before library preparation. For already fragmented DNA molecules (such as ChIP-DNA), the initial fragmentation step is omitted.

Stem-loop adapters are blunt end ligated to repaired input DNA. These molecules are extended then amplified to include barcodes using a high-fidelity polymerase to yield an indexed Illumina NGS library.

Library QC and sequencing

We evaluate the concentration of the library using qPCR and 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.

Last Updated: 3rd April 2024