Link to this post | posted 10 Jan, 2024 21:18

DanRussell

Hi Juan Carlos, Here in the Hatfull Lab at the University of Pittsburgh, all of our clinical M. abscessus isolates are classified as BSL-2, and for good reason. Sometimes an apparently "healthy human" may have undiagnosed or non-obvious issues, and treating an opportunistic pathogen as "harmless" is dangerous. As for the "ultimate, legal, foolproof" statement, that probably doesn't exist. BSL guidelines are set by your institution, and we at Pitt follow guidelines set by Pitt, the CDC, NIH, strain collections, etc. Different schools, countries, or organizations may have different rules, but we always expect all SEA-PHAGES schools to abide by appropriate BSL regulations in their region. We've had real safety concerns recently, as some SEA-PHAGES schools have tried phagehunting on bacteria they've isolated from the environment. While this offer a different phage yield, allowing students to work with an unknown bacterium and treating it as BSL-1 is not endorsed by the SEA-PHAGES program! –Dan Edited 10 Jan, 2024 21:37

Link to this post | posted 03 Nov, 2023 16:47

DanRussell

Hi SEA-PHAGES faculty folks, We got the following message and wanted to pass it on to you in case you're looking for potential summer employment. "The Summer Science Program recently designed a program in bacterial genomics where participants evolve a BSL1 bacteria (V. natriegens) to tolerate ever-increasing levels of antibiotics. After the evolution and Illumina sequencing our participants assemble the genome and try to identify putative beneficial mutations. We are looking for faculty to lead the micro and the bioinformatics sides, and Phage Hunters seemed like a good place to find faculty." Details on their employment page: https://summerscience.org/employment/ –Dan

Link to this post | posted 19 Jul, 2023 14:35

DanRussell

jcaoyao@gmail.com Hello Staff, I can't move files from my computer to the VM, so I tried clicking "Insert Guest Additions CD Image…", but nothing happened, no activity occurred in the terminal window or anywhere. What could I do, please? Thank you. Hi, you can try opening a terminal and running this command: sudo apt-get install virtualbox-guest-utils If that doesn't work, I'd suggest asking the internet! –Dan

Link to this post | posted 06 Jun, 2023 15:45
DanRussell	afreise Hi Dan - could we get a new forum for Cluster FP phages? Of course! Here you go: https://seaphages.org/forums/forum/222/ –Dan

Link to this post | posted 27 Jan, 2023 18:06

DanRussell

From Eric Miller: We are still recruiting undergraduates for the forthcoming summer 2023 REEU BeeMORE (Bees and Microbes) program at NC State University. Our goal is to recruit as many students underrepresented in agriculture and science as possible; our participants are selected based on their motivation to learn and to make the most of our 9 week program. Undergraduates in their second or third year are preferred, with most of them coming from outside the NC State University community. Here is the link to our home page: https://harvest.cals.ncsu.edu/beemore/ On the web page there is a recent video describing the program, a flyer suitable for printing or sharing electronically, and a link to the brief application procedure. We expect to keep the application window open for a bit longer. The program is funded for at least the next four years. Please share broadly! Thank you for helping get the word out, to your students and to your colleagues. Post it on one of your web pages citing summer opportunities! Thank you and all the best, Eric

Link to this post | posted 19 Jan, 2023 14:47
DanRussell	Hi Alison, Thanks. I asked because if they're the newer "M1" Macs, then we don't necessarily have a recommended way forward. (The chip architecture on the M1 Macs is different than the old Intel ones, and so VirtualBox will likely never work on M1s.) That said, if M1 is the issue, there's a big thread here on that: https://seaphages.org/forums/topic/5256/ –Dan

Link to this post | posted 18 Jan, 2023 16:29
DanRussell	Hi Alison, What kinds of computers do your students who are getting the error have? –Dan

Link to this post | posted 09 Nov, 2022 15:20

DanRussell

Hi Adam, 1. Yes, we use the standard Illumina protocol for denaturing/loading. The BioAnalyzer gives us numbers to adjust our starting pool to 4 nM. 2. Yes, we denature 5 µl of the 4nM pool. 3. We pool before we quantify, so we just take 5 µl of the pool. In your case, you'll need to pool libraries if you haven't done so. I'd recommend doing it in larger volumes than 0.25 µl though to avoid pipetting inaccuracy. If you've already normalized all the libraries @ 4 nM, then just take a microliter of each to make a 4nM pool, then take 5 µl out from that pool. (You should have plenty of extra library.) 4. We use the NEB Ultra II FS kit starting with 400 ng of input DNA. But other factors can determine final yield as well. For example, I think we only do 3 PCR cycles during the PCR step, but more cycles will get you more library in the end. Some libraries we've prepped are > 200 nM, others are only 5-10 nM. As long as you're above 4 nM, you should be fine. 5. Yes, we add between 3 and 6 µls of the phiX control before loading the sample. 6. Don't think so! Good luck, –Dan

Link to this post | posted 27 May, 2022 17:22

DanRussell

Hi Fernando, I'd recommend shooting for 100X coverage on your phage genomes. It might be a little higher or lower, but that's generally a decent target that's also not going to cost too much per phage. If you take an "average" phage genome of 70kb in length, that means you need ~7,000,000 bp of sequence to get to that coverage level. If the reads you're doing are 150 bases long, then: 7,000,000/150 = ~47,000 reads per genome Obviously that'll change depending on the size of the genome (often unknown!) and the length of the reads, so we usually shoot for ~100,000 reads per phage genome, though 50,000 will often be enough. As for protocols, we just use the standard protocol that comes with our library prep kit: https://www.neb.com/nebnext-ultra-ii-fns-dna/nebnext-ultra-ii-for-dna-library-prep After library prep, we use BioAnalyzer HS chips to check quantity/quality. Then the standard Illumina protocol to load the sequencer. We have a MiSeq, not a MiniSeq, so I don't have any specifics for the MiniSeq, but most of Illumina's sequencers these days are pretty easy to use. Take care and good luck! –Dan

Link to this post | posted 06 Apr, 2022 16:50

DanRussell

Hi Adam, Actually, they're not nearly nearly identical; there's only 32% amino acid identity between Arzan 58 and the next best match in the DB! (Which is Heisenberger 58 as of me writing this.) That's quite a lot of differences, and while it's probably reasonable to consider these homologs, they're still different enough that: 1. Phamerator's algorithm put them in different phams and 2. trying to make start calls on Arzan 58 based on Heisenberger 58 is not advisable. The AA sequences are 32% identical, but the nucleotide sequences show almost no similarity at all, so using a starterator report isn't a good idea in this situation, since SD sites and potential start codons have had lots of evolutionary time to change. This one you'll have to call on its own! –Dan