Link to this post | posted 08 May, 2018 19:12

kbutela

Hi Evan- This has happened to me on occasion while running the WINE version of DNA Master, most often if I have two files open at the same time and try to do a Frames view of both files. I'm guessing that some sort of memory issue may be at play here. Does this happen every time you try to work with this phage, and when this phage is the only .dnam5 file open in DNA Master? If you can send the file to me I can try to duplicate the error to see what's going on. Best, Kristen Evan Merkhofer Second error message.

Link to this post | posted 17 Aug, 2017 02:53
kbutela	Cluster B2s lack lysin B. I thought there was pretty good evidence of pham 609 being annotated as lysin B based on HHPred and TMM data. In this 2016 paper (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4912749/) they have Numberten and Pops (both B1s) containing both lysin A and lysin B.

Link to this post | posted 17 Aug, 2017 02:49

kbutela

In reviewing a few Cluster B1 phages, I am seeing some members of Pham (Webphamerator) 7880 (see Vivaldi gp64 for an example) as recombination directionality factor/RDF/RDF protein. I thought that the Cluster B phages are not temperate and aren't known to contain integrase genes. Why would a recombination directionality factor be present in a non-temperate genome, and apparently conserved among several other related phages? Is this a mis-annotation or am I not understanding something given that I'm writing this a little late at night? The only HHPred results don't meet the threshold for acceptance but they are close (high 80s%) to zinc finger domains. No conserved domains were identified. 114Kb

Link to this post | posted 11 Aug, 2017 21:39
kbutela	The documentation in the COILS2 help guide suggested a cutoff of 0.5 for a residue to be a part of a coiled coil motif. I may be a bit conservative in my interpretations, but I'm going to stick with the 0.5 cutoff for now. I attached a screenshot of the results I got when I ran Baka_5 through COILS2 using default parameters. 13Kb

Link to this post | posted 11 Aug, 2017 21:03

kbutela

Presence of a coiled coil motif. I ran Baka_5 as an example and very clearly saw the CCM Jackie identified (prob. was at least 0.6). Per our discussion on the Cluster A4 terminase genes from yesterday, I ran Blackmoor_2 through COILS2. Although this gene has HHPred hits (87% probability, 53% coverage) to a phage terminase, the probability of a CCM is only about 0.15. I played around with the scoring algorithms recommended in the manual and never got anything better. TinaFeyge_2 is annotated as a terminase, small subunit, but I also didn't get a CCM motif for this gene on COILS2. The evidence is close, but I don't think it meets the threshold to definitively call this gene in the A4s as a small terminase.

Link to this post | posted 08 Aug, 2017 20:02

kbutela

Pham 5142 (as of 8/8/17) corresponds to several members in Cluster B1, some of which are annotated as PLA2, PLA2-like domain, and PLA2-like domain protein, with PLA2 standing for phospholipase A2. According to this paper, some bacteria contain phospholipase A2, and a quick Blast search shows phospholipase A2 in Mycobacterium tuberculosis in two forms, one 512 aa long and the other designated as "membrane associated" at 111 aa long. Most members of Pham 5142 are relatively small, in the range of 40-50 aa. This gene (Virapocalypse gp73) does not match to any "PLA2" designated proteins in HHPred (attached) or in NCBI Blast other than mycobacteriophages. No conserved domains were found. I pulled the M. tuberculosis phospholipase A protein sequence (512 aa) and the one designated as "Membrane associated" as well as a sequence pulled from the crystal structure of phospholipase A from Streptomyces (1FAZ) and ran them through MUSCLE, Clustal Omega, and T-Coffee; all three give pretty lousy alignments with gp73 of Virapocalypse. I am not convinced this gene contains a PLA2 domain, but I'm not sure where the call of PLA2-like domain originated since the functional source for Virapocalpyse is ShiVal gp71 (older, annotated in 2011). Thoughts? 113Kb

Link to this post | posted 26 Apr, 2017 22:29

kbutela

I've been BLASTing just fine over the last few days. My DNA Master version is 5.23.1, build 2492, 29 January 2017. A lot of people, including myself, seemed to have trouble earlier in the semester, and I seem to recall that the group conclusion (maybe on another post?) was that the NCBI servers were getting too many simultaneous requests from the same place/school, so Blast was kicking people out, resulting in DNA Master throwing the error.

Link to this post | posted 02 Feb, 2017 22:21

kbutela

I'm running WINE DNA Master (29 Jan 2017 update) on a Mac OSX El Capitan 10.11.6, XQuartz version 2.7.6. I just ran autoannotation on some test files, and Glimmer appears to be working. Occasionally, I have encountered the Glimmer failure error, and just redoing the autoannotation later seemed to fix the problem. Keith's suggestion that the Glimmer servers may be getting clogged or rejecting multiple queries from the same IP domain in a limited amount of time may be a good explanation of what's going on.

Link to this post | posted 28 Jan, 2017 22:15

kbutela

I know there were some similar problems with the pre-2016 box file hosted at Baylor, but Tammy was able to get them to upload a version that worked, which is the current dmg file hosted on the Baylor Box site. Are they getting particular error messages? What exactly is happening when they attempt to open the file? Have they performed the update to the secure BLAST servers (some of my students forgot this step)?

Link to this post | posted 25 Oct, 2016 23:40

kbutela

If you are using DNA Master on the Mac via WINE and you've updated to Sierra OS 10.12, security changes to the Sierra MacOS may prevent you from opening DNA Master because it's an unverified program. Previously, this was solved by going to System Preferences > Security and Privacy; changing settings to "Allow apps downloaded from: Anywhere." The new Gatekeeper security features in Sierra have hidden this setting. Not to worry, as you can recover this "hidden" option by performing the following steps: Close "System Preferences" Open up a terminal window from your Utilities folder Type in the following command: sudo spctl –master-disable (Note: type in a double dash before the word "master" ) I'm not sure it's showing up well on this forum post Hit return and authenticate with your password When you reopen your System Preferences window, the "Allow apps downloaded from: Anywhere" will return as an option in the Security and Privacy window. In the long run, it may be worthwhile to add in an individual exception to DNA Master in individual Gatekeeper settings. However, I haven't tested this yet (instructions are here: http://osxdaily.com/2015/07/15/add-remove-gatekeeper-app-command-line-mac-os-x/). Will keep all of you posted when I figure out how to do this. Edited 25 Oct, 2016 23:41