GBS Guidelines and Pre-Submission Requirements:
Success in GBS is determined primarily by the quantity and quality of the submitted DNA. Please submit
the following quality control metrics with EACH project.
DNA Quantity:
A minimum 15 uL DNA at a concentration of 20-50 ng/ul is required for GBS. DNA should be RNAse
treated prior to submission. DNA should be measured by intercalating dye, for instance PicoGreen/Qubit
(Life Technologies) or Quantus (Promega). Nanodrop and other spectrophotometers are not acceptable
measures of DNA. We will re-quantify the DNA once we receive it using PicoGreen. For the purposes of
this assay, our QC measures are considered the gold standard, regardless of the quantity measured by
the lab. Lyophilized DNA is also acceptable, but we would prefer to have DNA suspended in 10mM Tris
or water. DNA concentrations do not need to be standardized across the plate; we will do this after they
are received.
DNA Quality:
To evaluate DNA quality, we require a gel image that includes a well-labeled ladder as well as an aliquot
of each DNA prep. Load 100ng of each DNA sample on to a 1% agarose gel, and run long enough that
the size standards are clearly separated. DNA should all be high-molecular weight (above the highest
size standard on the gel), with minimal smearing.
In order to test enzymatic activity, 10% of all samples submitted should be digested using a common
restriction enzyme that is NOT methylation sensitive (ex: HindIII or EcoRI). 300-500ng of each sample
should be digested for a minimum of 2 hrs at 37C, heat killed, then run on a 1% agarose gel as above.
Gel images should be sent to Marie Adams at marieadams@wisc.edu for approval prior to submitting
sample plates.
Optimization Plate:
If this is your first GBS experiment, please check to see if your species has been previously validated for
GBS by contacting Marie Adams at marieadams@wisc.edu. If it has not, we will need to create
optimized assay conditions for your species. To do this, we will need at least 10ug of DNA, either from a
single sample, or split between many samples. To optimize the species, we will test each set of samples
with several restriction enzymes and adapter concentrations in order to find a “best case scenario” for
your library. Optimized reactions create a library that has its highest concentration at 200-250 bp, with
fragments tapering off to 500 bp, minimal residual adapter contamination, and no visible indication of
repetitive DNA contamination (often visualized as a serrated effect on a Bioanalyzer chip).
Sample Submission:
Samples should be supplied to us in semi-skirted or non-skirted 96 well PCR plates. A good option is the
Eppendorf twin.tec semi-skirted plate (cat #: 951020303). Wells can be covered by either caps (ABGene
cat #AB-0783) or foil plate seals (Costar Cat #6570). Label each plate legibly with lab name, species
name, unique plate name, and date with a permanent marker. All plates must contain a blank well,
placed in a different random well in each plate. If partial plates are submitted, they will be treated like,
and charged as if they are, full plates. A sample submission sheet should accompany all submissions, and
can be found on our website.
Plates and submission sheet should be shipped on either dry or wet ice using an overnight courier to:
University of Wisconsin Biotechnology Center
Attention Marie Adams
425 Henry Mall, Room 1262
Madison, WI 53706
If you absolutely cannot meet these requirements, or have questions, please contact Marie Adams at
marieadams@wisc.edu for alternate arrangements.