FAQ – Frequently Asked Questions
1. How should my samples be labeled?
Samples should be labeled with your initials and a serial listing. For example, assume your name is Mary Lynn and you submit four samples. Your samples should be labeled ML1 through ML4. The next time you submit samples for sequence analysis, begin with ML5 and continue in this fashion. All tubes must be labeled individually.
2. When and where do I drop off my samples?
Samples can be dropped off Monday through Friday, between 9:00 am and 5:00 pm in Science, Rm 239. We recommend calling ahead to ensure a technician is present (208) 426-1038.
3. Do you archive/return samples?
Samples will be stored in the freezer for no longer than two months past the date of submission. DNA samples and sequencing libraries will be stored in a -20 degree C freezer. RNA samples are stored in a -80 C freezer. Please arrange for the prompt pickup of your samples.
3. How do I acknowledge your services?
Please support us by acknowledging our services in your publications.
Please add a sentence like this to your acknowledgments: “The sequencing was carried out at the Sequencing Core at the Boise State University.” Please email stephaniehudon@boisestate.edu when acknowledgments are made so we can keep track of publications that utilize our services.
4. What is your typical turnaround time?
Turnaround time is impacted by our sequencing queues. Our current turnaround time is approximately 1 week.
5. What buffer do you prefer for my samples?
We prefer you elute nucleic acids in nuclease-free water. Some buffers (such as TE Buffer) can interfere with downstream processes. If your samples are submitted in a buffer that is not compatible with downstream processes, we will either return the samples or do additional clean-ups to replace the buffer (at additional costs).
6. What are the causes of degraded RNA?
RNA is single-stranded and highly susceptible to degradation. Several measures should be considered when extracting RNA:
– Maintain an RNase-free environment. Wear gloves, designate an RNA-only workspace, and work quickly when extracting.
– Clean workspace with RNaseZAP – a cleaning solution that will remove RNase
– Use of RNase-free equipment, consumables, and reagents.
– Consider temperature. Extract from fresh tissue or quickly frozen tissue. Maintain RNA at -80ºC and avoid storing at room temperature.
7. What is the 260/280 ratio? 260/230 ratio?
Both of these ratios are obtained from Nanodrop and are used to determine sample purity.
260 = nucleic acid absorbance
280 = protein absorbance
The 260/280 ratio for pure DNA: ~1.8 and pure RNA: ~2.0.
230 = contaminant absorbance
The 260/230 ratio should be between 2.0 and 2.2.
If these values are lower, contamination should be ruled out. However, these ratios do not necessarily determine the success or failure of library preparations.
8. What QC is performed on the libraries?
Our typical library QC is Qubit and Fragment Analyzer analysis. All trace assays will provide base pair information for libraries which is used for normalization before sequencing. Unless there is a problem with your library preparation, we do not send final library QC results and will proceed with sequencing. Let us know if you would like to receive your QC documents.
9. How does Illumina sequencing work?
Illumina technology uses Sequencing by Synthesis (SBS). Learn more about Illumina sequencing technology by watching Illumina’s brief sequencing overview video.
10. What is the difference between single and paired end sequencing?
Single end sequences all fragments from one end while paired end sequences from both ends of a fragment. Paired end reads allow for increased mapping accuracy and isoform detection. Standard differential gene expression studies often require only single end reads. Paired end sequencing is beneficial when identifying genomic rearrangements, gene fusions, insertions, deletions, and repetitive sequences (among others). Learn more about Illumina paired-end sequencing.
11. Can you sequence libraries I created in my lab?
Yes. Please indicate the index sequences used in your preparation and if any custom sequencing primers are needed. All libraries submitted for sequencing will have QC to ensure the samples are ready for sequencing. While we do offer sequencing of user prepared libraries, we cannot guarantee sequencing success on library preparations performed outside of the Sequencing Core and you will be responsible for the cost regardless of the outcome. Please access the Illumina Adapter Sequences document to design libraries that are compatible with Illumina sequencers.
What is PhiX?
PhiX is a small, well-characterized bacteriophage genome. It consists of a well-balanced base composition with ~45% GC and ~55% AT. Because of this, it is used as an in-run control for run quality monitoring.
Learn more about PhiX.