Frequently Asked Questions
In this section, you will find some of the more frequently asked questions regarding sequencing reactions and data return. We hope this will be useful for day-to-day questions.
If you do not find the answer here, please browse our information pages or call 203-737-2566 or e-mail dnaseq@yale.edu.
- How do I ship my samples?
- Should I resuspend template in TE or H2O?
- How long is the average read?
- What size primers should I design?
- What Tm value should primers have?
- What should I do if I don’t have enough product?
- Why do I have to submit in a specific tube or plate type?
- Where can I get barcodes for my tubes?
- Can I leave empty wells in my high volume plate submission?
- Can I send less than 48 samples in a plate for a high volume submission?
- How can I make a RUSH sequencing request?
- Where are my results?
- Why does my sequence file contain NNNN?
- How do I ship my samples?
DNA samples can be shipped at room temp. If submitting in plates, we recommend sealing your plate very well (by using an adhesive heat seal or strip caps) and then wrapping the plate in parafilm. This should help to avoid any evaporation or leakage from well to well. Please ship to the following address:
Keck DNA Sequencing Lab
Attn: Jaime Heltke
300 George Street
2nd Floor, Room 2127
New Haven, CT 06511- Should I resuspend template in TE or H2O?
H2O is highly preferred. Sequencing reactions are very sensitive to chelating agents, such as EDTA.
- How long is the average read?
Using good template and primer, Taq/dye-terminator cycle sequencing will provide 500-600 bases of sequence with a 98-99% accuracy (exceptional template-primer combinations will yield 650-750 bases with 98% accuracy). We often see lengths of 800bp-1100bp. Template quality will affect length of read.
- What size primers should I design?
- Primers should be between 18-28 bases long.
- What Tm value should primers have?
Primers should fall within the range of Tm values 55-65oC.
- What should I do if I don’t have enough product?
You can submit half of the recommended reaction. We will still have enough product for 1 repeated reaction, if necessary. Just make sure everything is in proportion to the original reaction.
- Why do I have to submit in a specific tube or plate type?
- We use automation for our processing, so any incompatible plasticware results in longer turnaround time, extra labor, and associated transfer fees. To avoid these issues, we recommend you adhere to our requirements.
- Where can I get barcodes for my tubes?
- Barcodes are given out over the counter in the Kline and Medical Stockrooms. You can also find them outside of S340 TAC, BCMM 108, and our lab at 300 George Street, Room 2127 (come visit us!).
- Can I leave empty wells in my high volume plate submission?
- You may leave empty wells, but we charge for every well in between A01 and your last well with sample. Please remember to fill sequencing plates by row. Only fragment analysis plates should be filled by column.
- Can I send less than 48 samples in a plate for a high volume submission?
- Yes, you may. It is actually more cost-effective to use our high volume service for sequencing if you are submitting 25 or more samples. We urge you to use this service if you are experienced. It saves on time by eliminating the need for barcodes and excessive tube labeling.
- How can I make a RUSH sequencing request?
- Please email us before we receive your submission. We process all sequencing samples in batches of 96, and will work to accommodate your request without compromising the lab’s workflow requirements. If your request is a fragment analysis plate or for our QC service, we will get it processing as soon as we are able.
- Where are my results?
- Your results are uploaded to Box.com and should be available in 1 business day after receiving your samples. Please make sure you have accepted the invite to Box.com (check your junk mail). If you have not received your results within 2 business days, feel free to contact us at DNAseq@Yale.edu.
- Why does my sequence file contain NNNN?
- There is an insufficient level of termination products for the computer software to call bases. There is no usable data to report. Please refer to our Troubleshooting Help page for more in-depth information.