UNIVERSITY OF COLOGNE

For whole genome sequencing (WGS) we routinely use PCR-free and PCR based protocols. Options for FFPE samples are also available. For a human genome with 30X coverage we sequence 120 Gigabases. The standard read length is 2x150bp. Data analysis for human samples is possible through our Varbank genome pipeline (germline variants).

Long read workflows are available for both Oxford Nanopore and PacBio technologies. Frequent applications for long read data are de novo genome assembly or structural variant calling. The read quality has drastically improved in recent times, so that polishing with short reads is commonly not necessary. Sequencing is possible on the latest MinION/PromethION and Revio flowcells. Sequencing of HiFi libraries on the Revio system is done in cooperation with the WGGC in Düsseldorf.

We use the Agilent SureSelect enrichment kits (v8 for human samples) and the XTHS2 library protocol with UMIs. This is usually applied to human and mouse samples, but other species may be possible on request. FFPE samples can be submitted, but performance may vary depending on sample quality. Data analysis for human samples is possible through our Varbank exome pipeline (germline variants).

We use the enzymatic conversion method (EM-Seq). We include methylated and unmethylated controls as spike-ins in the library preparation, and check the conversion rate for these controls after sequencing. Sequencing read length is 2x150bp. We have also run this protocol on low input samples (cfDNA).

You need to provide the purified amplicons. We can prepare the library and sequence it 2x100bp or 2x150bp on the NovaSeq. In this setup you can freely choose the number of reads you need for your amplicon sample. Sequencing on the MiSeq is also possible with up to 2x300bp read length. For this option your sample will be sequenced on one complete MiSeq flowcell. Please be aware that in this case the complexity of your sample has an impact on the sequencing output. Full length 16S amplicon library preparation and sequencing (~1500bp) is also possible (Nanopore).

You need to provide the purified DNA after the transposase reaction in a volume of 20µl (elution in water or 10mM Tris pH8). We will use the entire input volume to prepare the library. We will not do an initial quality control of the input material. Sequencing is possible with 2x100bp or 2x150bp.

You need to provide the fragmented and purified DNA after immunoprecipitation. Please specify which protocol you are using (Cut&Run, Cut&Tag). Provide the sample in a volume of 20µl (elution in water or 10mM Tris pH8). We will use the entire input volume to prepare the library. We will not do an initial quality control of the input material. Sequencing is possible with 2x100bp or 2x150bp.

Please contact us for the available protocol options. We start with the crosslinked (and optionally biotinylated) DNA fragments.