Quality Control

QUALITY CONTROL, PURIFICATION, AND SIZE SELECTION

SERVICE DESCRIPTION

We offer quality control services that include initial QC for library development, size selection, and final QC of libraries that have been prepared by you and are ready for sequencing. Our scientists have years of experience analyzing and libraries and troubleshooting issues. We use state-of-the-art equipment for quantification and sizing of DNA, from initial library input to finished library. Accurate QC ensures high quality libraries are run on the sequencers at the same concentration and helps identify adapter/primer dimers and unintended products that may cause sequencing issues to arise. We also offer automated DNA purification and size selection prior to or after library preparation for precise, discrete selection of DNA.

 

SERVICE OPTIONS

Standard DNA Quality Control

Our standard DNA QC service includes fluorometric quantification and size analysis on the Agilent 4200 TapeStation System of your DNA or Illumina libraries. This service is ideal for shorter DNA fragments (e.g., sheared, amplified, digested, Illumina libraries) of 35 bp to 1,000 bp and concentrations as low as 5 pg/µl. Quantification is performed first to inform us which TapeStation assay is best suited for your samples. This service is required for all Illumina libraries submitted for sequencing.

High Molecular Weight DNA Quality Control

Our high molecular weight DNA QC service includes purity analysis on the NanoDrop One or Synergy H1, fluorometric quantification, and size analysis on the Agilent Femto Pulse, an automated pulsed-field gel electrophoresis system. This service is ideal for larger DNA (e.g., genomic, linearized plasmids) of 1.3kbp to 165kbp+ and is required for all DNA being submitted for Oxford Nanopore and PacBio library preparation and sequencing. This service can also be requested when higher quality assurance is needed, regardless of size.

Solid Phase Reverse Immobilization (SPRI) Bead Purification

SPRI bead purification is an option for libraries containing adapter, primer dimer, and/or small, undesired fragments that may cause issues during sequencing. Libraries with ≥ 1% adapter dimer, by molarity, cannot be sequenced on the NovaSeq and libraries with ≥ 5% adapter dimer cannot be sequenced on the MiSeq. SPRI bead purification is performed using Axygen™ AxyPrep Mag™ PCR Clean-up beads (Corning Inc.). Following SPRI purification, QC is performed again to ensure effective purification. We offer SPRI purification in situations where DNA purity is compromised by organic compounds or other contaminants/inhibitors. We also offer this service for cleanup of reactions (e.g., PCR, enzyme digest) as the process is automatable and high throughput.

Size Selection

We perform size selection of DNA or prepared libraries on the PippinHT and BluePippin (Sage Science). These platforms provide medium throughput size selection. PippinHT allows for precise size selection of up to 20 ul of DNA or prepared Illumina library.

 

LIBRARY QC WORKFLOW

Access our Interactive Submission Guidelines and get the exact guidance you need to prepare, label, and deliver your samples.

 

If you can’t find the information you’re looking for, please contact us at nextgen-seq@biotech.wisc.edu

Standard and High Molecular Weight DNA Quality Control Pricing

Effective 12/1/21

* Quantification and size analysis of Illumina libraries and other DNA. Cost per sample, based on batch size.

SPRI Bead Cleanup and Size Selection Pricing

Effective 12/1/21

* Libraries or other DNA that undergo SPRI bead cleanup will subsequently incur a QC charge according to the Standard DNA QC pricing, in addition to the initial QC charge.

Unless otherwise specified, all data and reagents distributed by the University of Wisconsin Biotechnology Center DNA Sequencing Facility are intended for research purposes only. They are not intended nor certified for diagnostic or clinical use. Clinical services are provided through our collaboration with the UW Collaborative Genomics Core.