Genomic Technologies Facility › Frequently Asked Questions (FAQ)
2025 Roy J. Carver Co-Laboratory, Iowa State University
1111 WOI Road, Ames, Iowa 50011-1085
Tel: (515) 294-7491 | Fax: (515) 294-5256 | Email:
Ion Torrent Proton [ Top ]
How much library should I send and at what concentration?
Please submit between 10-20 µL of library, it should be at least 500 pM. The molarity of the library can be calculated by qPCR using KAPALibrary Quantification Kit or by Bioanalyzer DNA High Sensitivity chip or DNA 1000 chip. Samples should be submitted in 10 mM Trus-HCl or Nuclease-free water. Please GTF staff if your library's concentration is lower than 500 pM.
Do I need to provide any quality scores or Bioanalyzer reports?
Please send the Bioanalyzer ".xad" file to when you are ready to submit your sample. We use this file to check your DNA quality, concentration, fragment length and check for any short DNA fragments in your library that would inhibit or reduce the ability of your library to template. Please include the dilution factor if you had to dilute your library before running your sample through the Bioanalyzer.
Does GTF provide library quantification services?
Yes, we will quantify libraries using your choice of the Agilent 2100 Bioanalyzer, or via RT_PCR using the KAPA Illumina library quantification kit. For certified users, the KAPA kit reagents are also available for use. Fees for these services are not included in library preparation or NGS runs.
Do I need to use a specific library kit to prepare my library?
No. The only requirement is that your adapters are the Ion A and P1 adapters. They are available from Life Technologies or you can choose to have them synthesized by your preferred oligo synthesis company.
Is there a size limit for the insert?
Ion Proton Sequencing is limited to a maximum insert size of 200 bp. With adapters, the maximum fragment size should not exceed 260 bp.
How long does a sequencing run take from library delivery to data?
Ion Torrent sequencing has 4 steps to generate data. The first three steps involves 8 hours of template preparation, followed by a maximum of 4.5 hours for sequencing.
- Template generation: emulsion PCR to amplify library sequences on Ion Sphere Particles (ISPs).
- Template QC: a test to ensure enough library successfully template on ISPs.
- Enrichment: a purification step to remove non-templated ISPs and library sequences not adhered to ISPs.
- Sequencing: loading of ISPs into an Ion chip and sequencing run.
How many reads can I expect from a sequencing run?
The Proton P1 chip generally produces 80-100 million reads per run.
What are the steps leading up to sequencing for Ion Torrent technology?
After reviewing your Bioanalzyer report, we will dilute your library for template preparation, template your library onto Ion Sphere Particles (ISPs), enrich the ISPs for library templated ISPs only, load ISPs into the desired chip, and sequence.
How can I get my sequencing data?
The GTF can send your data via FTP services or you can choose to send a hard drive or large flash drive to the GTF and we will download the data onto your storage device of choice and send it back to you. We recommend you back up your data immediately upon receipt, because the GTF will be unable to store sequencing data beyond two months due to the large size of the data files.
Qiagen BioSprint 96 [ Top ]
Can the BioSprint 96 be used to extract DNA from non-plant species?
GTF stocks the Qiagen BioSprint DNA Plant Kit. Other species may require a different kit. Please contact Qiagen to determine the kit that will fit your needs and provide your own kit if using something other than the DNA Plant kit.
How long is each run?
Each run will take about 30 minutes to complete. There are several plates that must be prepared prior to a run; GTF will have most of these plates completed before you arrive. Thus, hands on time will include preparing your sample plate and loading the machine for each run.
What is the required input?
For maize, Qiagen recommends 50 mg of fresh leaf tissue, or 10-30 mg of dried tissue. For other species, please refer to Qiagen for recommendations (http://www.qiagen.com/us/products/catalog/sample-technologies/)
How should I provide samples to GTF
Samples should be provided as freshly ground tissue. Please contact GTF staff to discuss extra charges if you are in need of additional services for drying and grinding samples.
Is it required to have 96 samples?
No, you can do a run with fewer samples. However, GTF charges are based on the number of plates used.
Do you provide quantification services after my DNAs are extracted?
Quantification services are not included in the cost of extraction. For an additional fee, GTF can quantify samples using the Eppendorf AF2200 Plate Reader.
I see some black particles precipitated in some wells. What are those? Will they affect my experiment?
Black particles that can be seen in the wells are leftover magnetic beads used during the extraction protocol. According to the Qiagen manual, these beads will not affect downstream applications. However, if you wish to remove them, simply place your plate on a magnetic block and transfer the elution to a new plate.
Eppendorf AF2200 Plate Reader [ Top ]
What kind of quantification measurement methods are offered?
The Plate reader is capable of taking both absorbance and fluorescence based readings. For absorbance, readings can be taken in a plate format, or by using the Eppendorf µPlate 0.5, which can hold up to 16 samples at one time.
How long does it take to quantify 96 samples in a plate?
It takes about 15-20 minutes to quantify 96 samples.
What types of plates can be used?
The type of plate to be used depends upon the application. For quantifying small volumes (2-4 µL) GTF provides the Eppendorf µPlate 0.5, which can hold up to 16 samples at one time. A variety of 96 well plates can also be used. The use of 96 well plates requires a sample volume of 100 µL. Fluorescence based assays require a black plate. For a full list of acceptable plates, please refer to the operating manual at http://www.eppendorf.com/int/.
How do the absorbance measurements from the Plate reader compare to those of the nanodrop?
While the measurements are not exactly the same, there is a very high correlation between Plate Reader and nanodrop sample readings. Given the slight discrepancy, GTF recommends that all samples from 1 given project be quantified using the same method.
How can I have my samples quantified?
GTF offers the Plate reader for on-campus user at no charge. On-campus users must provide their own consumables and use their own pipettes. GTF staff can quantify samples at cost (labor and consumables, dependent upon the assay type).
Eppendorf epMotion M5073 [ Top ]
Which tools does GTF provide?
GTF currently offers the 50 µL and 1000 µL dispensing tools.
What do I need to do to use the epMotion?
GTF provides the epMotion for certified users at no charge. A training session is required to become certified. Users must provide their own epTIPS, which can be purchased from the facility at a minimal fee per box.
General Questions [ Top ]
What are GTF hours?
GTF is open Monday through Friday from 8 am to 5 pm.
Where should I send my samples?
Mail your samples to:
Genomic Technologies Facility Manager
Iowa State University
2025 Roy J. Carver Co-Laboratory
1111 WOI Road
Ames, IA 50011-3650
Any special instructions for shipping?
If you are shipping overnight or 2nd day, ice packs should be sufficient. If you are going to ship over a longer time period, you should use dry ice. Insulated containers are best.
How should I ship my samples?
For small sample sizes, we recommend 1.5 or 1.7 mL microfuge tubes, wrapped with parafilm. For large genotyping experiments, place DNA in 96 or 384-well plates, sealed with tape.
When will bills be sent?
Bills are processed at the end of each month. Invoices are mailed to the billing address on the submitted User Agreement.
Can I pay with a credit card?
No, GTF no longer accepts payment via credit card. Other options for off-campus customer payments include wire transfer or check.
What are the terms and conditions for using GTF services?
Please refer to GTF Terms and Agreements. Please indicate that you have read these terms on the user agreement form submitted before beginning a new experiment.
What happens if my experiment does not work?
GTF strives to provide high quality services and to achieve customer satisfaction. We want to ensure our clients that we are here to provide our best services. As a non-profit service center, we provide service at minimum operation costs. In the event of poor quality results, we will work with our clients to determine what the possible causes may be. If the cause(s) is from operation at GTF (i.e. software or equipment malfunctions), GTF will cover 100% of the cost of the failed portion of the experiment. If the cause(s) is from the client-submitted samples (i.e. low concentration or poor quality of samples) or incorrect information provided to GTF, the customer will be charged for the failed run. If a cause cannot be determined, GTF and the customer will split the cost equally. Please if you have any questions or need additional information.
How can I become a certified user?
GTF allows certified Iowa State University users to operate several instruments themselves. A one-hour training session is required for use of the Mx4000 and LC480 RT-PCR machines, Blue Pippin, Bioanalyzer, and E-gel size select system. Users will be required to provide their own consumables (pipet tips, gloves, etc.).
Customers wanting to use the Proton or the MiSeq will need to undergo several extensive training sessions to become certified. These sessions involve shadowing GTF staff as they perform at least two experiments (~6 hours each), learning how to properly run and clean the instruments, and having GTF staff supervise your first independent run. GTF staff reserve the right to refuse certification to any individual. Expect each training session to last at least one full day. For example, the workflow for one Proton run is:
7-8 am: set up one touch amplification
8-12 pm: 4 hour break
12-1pm: prepare chip
1-2 pm: start cleaning and initialization
2-6 pm: wash one touch, enrichment, QC, start run
Training sessions are charged at $35.15 per hour. Please contact GTF staff at to make arrangements for training sessions. Certified users must submit a user agreement before training sessions and fill out a usage form for every experiment. Certified users accept responsibility for all experiments and abnormal equipment breakage.