Link to this post | posted 11 Jul, 2024 21:10
kmaclea	I am glad to see this advice as it jibes completely with my experience! – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 12 Jan, 2024 15:52
kmaclea	Thank you David for your long service to HHMI and the SEA! We are truly grateful for your work! – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 03 Oct, 2023 18:22

kmaclea

Bernadine and Adam I had to reset my password, so thanks to Adam for sharing what I had sent him on email. Yes, it's clear that least in our old mill building we have some serious thermophile (or thermophile spore) contamination. Since this is the only class we see it, but also the only class where we had commonly held anything at 55C for any length of time, it tracks. There could be other issues above that were separate issues, but our main continuing issue seems to be the thermophile, and our immediate solution to it as has been the single-use top agar, and only either making it fresh or melting it only right before use out of the fridge. We have had a separate issue with making sure that A. glob. 2979 or 2880 are healthy and uncontaminated, so we have become very paranoid and rigorous in our handling, making single-use host cultures as well. Thus far, our single-use approach has been working, and quickly got us back to numerous plaques on direct isolation with A. glob. 2979. If you have any further questions, please reach out! Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 03 Mar, 2023 19:29
kmaclea	Just a note that we continue to find this in working on MulchSalad (F1) genes. We saw some genes today in which RefSeq is calling the gene one thing and INSDC is instead calling it a hypothetical protein! So it even gets more complicated! Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 24 Feb, 2023 20:52

kmaclea

So we've got another instance of the same issue, Debbie. MulchSalad_Draft_37 is an integrase/tyrosine integrase by analysis on Phagesdb BLAST/HHPred as shown in the pham report: https://capture.dropbox.com/TobVF9m4QnDFz6Hl NCBI BLAST is showing mostly "endonuclease" as the annotation: https://capture.dropbox.com/g0O9azXO34T0rhG9 And if you do the "Identical Proteins" analysis on any of the predicted "endonuclease" genes you see the difference again: https://capture.dropbox.com/RTPOWS6fziZ4TMpK What we're seeing again is the RefSeq ("curated" database is calling most of our Cluster F pham 68632 integrases as endonucleases whereas GenBank/INSDC and Phagesdb are calling them integrases. So somewhere along the way RefSeq "curation" is changing the default of what we are calling these genes. And since apparently, by default, NCBI BLAST shows the RefSeq results and only shows the GenBank results if you dig deeper, I'm guessing this will cause a certain amount of confusion going forward. Meh. Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129 Edited 24 Feb, 2023 20:54

Link to this post | posted 23 Feb, 2023 17:33
kmaclea	Thank you–I like to use the MTPs as an example and it's nice to be reminded of the parameters around the function call! (This silly RefSeq issue notwithstanding!) – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 19 Feb, 2023 13:21
kmaclea	That is very helpful, Debbie! Thank you! My inclination was to ignore this particular NCBI RefSeq call–so I am glad to hear you agree. And in the meantime I am definitely hoping this isn't more widespread! Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 17 Feb, 2023 19:50

kmaclea

I start out this post with the caveat that I may be doing something really dumb here, so I apologize if this is a known issue and I have just somehow missed it. Here's the situation: We're annotating MulchSalad (F) and had just started to teach function calling. We picked gene 1 to demonstrate with (big mistake! ). Basically here's the issue: If you BLAST on Phagesdb this gene hits to terminase small subunit. This is shown in the Pham view for other annotated genomes. If we BLAST (blastp) on NCBI with default settings, we see "minor tail protein" for the annotation: https://capture.dropbox.com/ZrKNG9p6b1vT2pDa I was confused by this and dug deeper. Apparently, this is because RefSeq is defaulting for matches. INSDC regular GenBank entries are still there with the names given by SEA-PHAGES, but they are hidden in the hits by default. You can directly compare this if you click around: https://capture.dropbox.com/u8NpcBuUFOAcg2UT If you look at the RefSeq entry it gives "minor tail protein" and if you look at GenBank it says "terminase small subunit" I am not sure if this is a common problem, but we definitely found it here. I am not sure there is really a question here so much as an observation. If I *do* have a question it's likely unanswerable–why did RefSeq call this MTP? HHPred agrees this is a terminase large subunit or terminase. Is this a common issue? This is the first time I'd ever seen anything like this. If it's isolated, I can deal with it ad hoc… if it's common maybe I need to plan a workaround. Thanks, all! Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 09 Feb, 2022 16:53

kmaclea

I was having Mary's error too on a new VM. I took note of Dan's message I had no apostrophe in my path and had placed the file in a \DNA Master folder on my desktop (which had a space in there). I decided to make things very simple. I created a C:\DNAMaster folder. I put the installation program there. I ran the installer (which was in my Downloads folder) as an administrator with that folder as the destination (and the installer wanted by default to put the program in C:\DNAMaster\DNA Master (again with the space)–but I changed it just to C:\DNAMaster and installed. Then I ran DNAmas.exe from that folder, also as an administrator. I did not the get the same errors as I did the first time (which are just like Mary's). Instead I got: Field error (then) Field error (then) Data Module Error creating GenomeClipboard.db, tables Then I updated the program. And then, it is now seeming to operate normally. Posting this in case it is helpful to others. I am not sure if the space character in any part of my path was at fault, but I had no apostrophe or other special character in there, so I decided to try this as a last resort before asking for more help. But since it seemed to work–I'm going with it. Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129

Link to this post | posted 14 Jan, 2022 22:20

kmaclea

I'm still a little unsure how to turn this into Notes that QC reviewers downstream can use once we submit our genomes. Are we suggesting that one will submit the DNA Master Minimal File, and then an attached Spreadsheet or linked Google Doc that has info on each gene, which will then be reviewed alongside the Minimal File by the QC reviewer? And that the files in question do not have to follow any particular format? So some schools may submit much larger files than others? Are there guidelines on length or what is included (screenshots, etc.)–because a spreadsheet can compactly give certain info, but a Google Doc with inserted screenshots, etc., would be very different. Thoughts from anyone? Kyle – Kyle MacLea Associate Professor, University of New Hampshire at Manchester kyle.maclea@unh.edu +1 603-641-4129