It's been a weird year, but through many trials and adjustments we have still arrived at phage DNA sequencing season! This post contains information about sending your SEA-PHAGES phage DNA samples to the University of Pittsburgh for sequencing. Because of the pandemic and other issues, several things have changed since previous years, so please read carefully.

Contact Us/Questions

Please send any questions about the information below to Dan (dar78@pitt.edu) and Becky (rag4@pitt.edu).

Sample Submission Form

Please submit the information about the samples you are sending using Google Form. This will help us keep track of samples, correct any spelling errors, and make sure your samples are accounted for. Please fill out a copy of this form for each sample you are submitting. The best time to fill it out is as you are getting your shipment packed, so that when your box arrives, we'll already have the information on your phages in our database.

NEW THIS YEAR: We are asking, if possible, that you add a tracking number in the appropriate place on the form. We have rotating lab staff, and want to make sure all packages are accounted for. Please note whether each sample is a "Priority" or "Backup" in the Notes field, along with any other info we should have.

https://docs.google.com/forms/d/e/1FAIpQLSfdhZpw1LHL42zxAotEVqhmrr8RBN9-kuPL_42ioYmuBU-4-Q/viewform

Quantity

Each SEA-PHAGES school may have two genomes sequenced per Bioinformatics section that it is teaching during the current academic year. Please indicate any order of preference, if you have one.

NEW THIS YEAR: Because many schools were not able to perform phage discovery as usual during the fall but still want to do bioinformatics in the spring, we will need some extra DNA samples to be donated to the Genome Exchange. If you have managed to extract more good quantity, high quality DNA samples than you need for your class, we would welcome those as well. Please indicate "Donate to Exchange" on the Notes field of the form. And thanks for helping out your fellow SEA-PHAGES institutions!

Deadlines

Semester Schools

Please send your genomic DNA so that it arrives in Pittsburgh by Friday, November 20.

Quarter Schools

Please send your genomic DNA so that it arrives in Pittsburgh by Friday, January 8.

If you will have trouble meeting these deadlines for any reason, please contact us.

Guidelines for DNA

Buffer

Samples that are resuspended in TE are problematic, since the EDTA may interfere with enzymatic shearing of the DNA. You can resuspend your DNA in Elution Buffer (Tris) or in purified water.

Amount

We need a minimum of 4 µg (micrograms) of genomic DNA. If you're in the ballpark of this number but not quite there, contact Dan and Becky to see if it's okay.

Concentration

At least 40 ng/µl. Ideal is 100-300 ng/µl. If you're above 300 ng/µl please consider diluting to a workable concentration and workable volume. Shipping volumes less than 20 µl is not recommended. Keep in mind that spec-based quantifications (like Nanodrop) count all absorbance at 260 nm, not just genomic DNA, so they often overestimate the real concentration. Err on the high side of the amount of DNA you send.

Gel Picture

Please submit—either via email or in the shipping box—an image of the gDNA being run on an agarose gel. We use these to look for sample integrity (not already sheared/degraded) and purity (no RNA).

Labeling

Please clearly label the tubes you are sending with the name of the phage as it appears on PhagesDB. Don't label tubes using student's initials only, or other ambiguous names like "Phage1" or "PittPhage". If possible, put a small circular sticker on the top of the tube and write the phage name only on it.

Shipping

Packaging

We prefer standard microcentrifuge tubes, and strongly recommend wrapping the caps with Parafilm to prevent spilling or evaporation. The tubes can be packed into a 50 ml conical tube with some KimWipes to stabilize them during shipment. If shipping overnight (preferred), you can actually just send the samples at room temperature, as the DNA should be fine for 24 hours in transit. If shipping on a slower schedule, you should use cold packs. Aim for delivery to Pittsburgh on Tuesday-Friday. If shipping near holidays, please check with us about when we'll be around to receive samples.

You can include a packing slip with any information you think we should have about the enclosed phages.

Address (NEW THIS YEAR)

Attn: Becky Garlena
University of Pittsburgh
Langley Garage 120F
Langley Hall
Fifth and Ruskin Avenues
Pittsburgh, PA 15260
(412) 624 6976

Additional Samples

For the past several years, the Genomics Sciences Laboratory at NC State has provided high-quality, for-cost sequencing services to schools in the SEA-PHAGES program who wish to sequence additional genomes beyond those allotted by the program. Contact Andy Baltzegar at dabaltze@ncsu.edu for pricing and details.

Archiving Samples

Just a note that archiving samples should be sent to the same address as above, and so if it's convenient to you, you can send DNA samples and archiving samples together in a single box.