DNA Sequencing

Due to new NIH policy, all users should acknowledge any Core used in the actual Acknowledgements section. This is due to the use of an automated check of all the Acknowledgements section to determine what resources were used for the publication. This information is used to impact grant funding for the entire institution. Therefore, please be sure to acknowledge us or any other Cores you use for your publications.

Please put the following in your acknowledgements section:

Sequencing was performed at the DNA Sequencing Core Facility, University of Utah.

Thank you.

The DNA Sequencing Core will not retain researcher’s data after delivery of the data to the requesting researcher.

Per NIH policy, it is the responsibility of the grant holder to store their data for at least the minimum time required by the grant (usually at least 3 years, but please check your grant conditions).

Illumina sequencing available. We are able to do Illumina sequencing through an off campus contract. Please read here for for more information.

Standard Sanger Sequencing is commonly used to determine sequence of PCR products, verify plasmid inserts, genotype samples, and validate Next Generation Sequencing results.

We provide support ranging from primer design and Sanger validation for labs through simple running of samples submitted by labs. We spend many hours every year helping labs troubleshoot and plan projects.

Often Used For:

• Plasmid insert validation
  Many research techniques require cloning of regions of interest into plasmids for further research. Once insert is inserted into the plasmid, a method of validating that the correct sequence was obtained is necessary. Sanger is usually how this is done.
• PCR sequencing
  Often research requires the interrogation of distinct sequences within organisms. If sufficiently stringent primer sets can be designed, then PCR amplification and subsequent Sanger Sequencing is often the method of choice. Also frequently used to validate putative mutations discovered by NGS.

Drop-Off LocationsHow-To Guide: Submitting an OrderHow-To Guide: Looking at Your DataPrimer SequencesSample Submission GuidelinesSample TurnaroundSequencing PricingSequencing Software

Sanger Validation of NGS Data     

$50/hour time and any reagents for testing (primers, PCR mixes, gels, etc)

*BACs/large Clones; Talk to us about Ion Torrent

Next Generation Sequencing (NGS) is rapidly replacing Sagner Sequencing and other methods as prices continue to drop and instrumentation continues to improve. We own and operate an Ion Torrent Proton.This platform provides options to obtain sequencing quickly and without needing large numbers of samples. This platform is particularly well suited to small genomes and targeted sequencing.

Pyrosequencing is often used to interrogate methylation states of DNA and genotype small numbers of SNPs across sample cohorts. It is capable of sequencing approximately 150bp off of known adapters.

The pyrosequencer is offered for use either directly by the laboratory (once trained) or as a complete service run by the core. For scheduling use of this instrument, please email labtech@cores.utah.edu or use our scheduling system. Runs must be scheduled in advance with the Core.

This is cost per run to cover service contracts. This does not include costs of your kits that will be specific to your project and will need to be priced with each project.

Pricing for runs performed by the core: Please Inquire by email  labtech@cores.utah.edu.

Labs will be responsible for purchasing their own reagents and consumables. The Core can help with part numbers and pricing through Qiagen.

The DNA Sequencing Core has a Beckman BioMek FX which can be used to aliquot samples from plates or tubes to other plates. If this is something that would help your group, please contact the core to discuss further. Robot time can be signed up for in the scheduling system, BUT PLEASE CONTACT US BEFORE ASSUMING THE TIME WILL BE FINE. If this is a new project, come talk to Derek before proceeding as it takes time to design and test protocols to meet your needs.

Pricing: 1/4 hour increments at $60/hour plus any consumables

The NIH has added requirements for all grant holders to authenticate key biological or chemical resources.  (http://grants.nih.gov/grants/guide/notice-files/NOT-OD-15-103.html)  This includes authenticating any cell lines that you may use in the research.  Many journals are also now requiring this.  Cell line misidentification has been an ongoing problem is science for many years.  Two surveys estimate cell line contamination to be between 20-36%.

We here at the DNA Sequencing Core want to be sure our customers are able to publish not only correct data, but in any journal they would like.  Therefore we have introduced a cell line authentication service.  We offer testing at 24 loci to determine the genotype of your cell line with certainty.  The current standard is 8-10 loci per cell line.  Although this is theoretically sufficient to separate different cell lines, it is not necessarily sufficient if the cell lines are related.  For example, we did initial testing of Jurkat and JLAT cell lines.  These two cell lines were identical across 20 loci (including the standard 8-10 currently used), but differed at 4 loci.  JLAT cells are derived from Jurkat cells.  Understanding how closely related these two cell lines are illustrates the importance of being able to test at 24 loci instead of the current 8-10 loci standard.

No database currently exists that contains all 24 loci, so initially matching of more than 8-10 loci will be difficult.  We recognize this is a problem and have, therefore, joined with ABRF to test cell lines obtained directly from cell line providers across the 24 loci we test for.  This data will be entered into a database that we will house here at the University of Utah, but which will be publicly available for any researcher wishing to access it.  This database will be small to begin with, but will continue to grow as we are able to test more and more cell lines.

We invite you to contact us to submit your samples for this service.  Runs will initially be once a week and we will consider adding more runs as necessary.  Initial pricing will be as follows:

For more information, feel free to read more on Cell Line Authentication from the manufacturer of the kit we use.

https://www.promega.com/products/cell-authentication-sample-identification/cell-line-authentication/cell-line-authentication-testing/

The Fragment Analyzer system is intended for quantifying and qualifying checks of DNA, gDNA, and RNA samples.  The system runs various kits measuring a range of fragment sizes from small PCR amplicons to very large DNA (10bp – 40,000bp).  The system has been adopted and incorporated into many current workflows, including for Illumina and PacBio NGS systems for library QC and input sample validation.

The system detects nucleic acids by using an intercalating dye and then running the samples out on a gel matrix in a very similar fashion to a capillary sequencer.  Results can be used to determine DNA or RNA quality before moving to sensitive downstream applications like NGS libraries, array runs, etc.

Results are delivered to the customer in the form of a PDF report from the manufacturer’s ProSize software.

More information about the Fragement Analyzer is available on AATI’s website here:  https://www.aati-us.com/instruments/fragment-analyzer/

Costs per run can vary by kit, but will be very similar.  One run is 1-11 samples plus a ladder.  The instrument runs all 12 positions whether or not there is sample.  Input amounts vary quite a bit kit by kit.  Please contact the core to discuss sample concentration.  We will use 2ul of the appropriate concentration per sample.  The kits currently stocked by the core:

High Sensitivity NGS Fragment Analysis Kit:

High Sensitivity RNA Analysis kit

Monday-Friday 9am-5pm excluding University closure days

Radiobiology Building (Building 585)
30 N. 2030 E. Room 110 RB
Salt Lake City, UT 84112

DNASequence@cores.utah.edu
Labtech@genetics.utah.edu

801-585-2976 – DNA Sequencing Core
801-213-2928 – Billing and Accounting
801-585-2978 – Fax

Derek Warner, Director (Hablo Español)
801-581-4736
dwarner@cores.utah.edu

Michael Powers, Sr. Lab Specialist
Phone: 801-585-2976
michael.powers@cores.utah.edu

Lynn Jorde, Ph.D., Professor Human Genetics

Colin Dale, Ph.D., Professor, Biology

Robert Weiss, Ph.D., Professor, Human Genetics

Aaron Quinlan, Ph.D., Professor Human Genetics