Link to this post | posted 10 Mar, 2025 18:31

debbie

Hi Christy, Yep. It looks like the DNA Master website is down. (I've made an inquiry.) You can find the program that is needed in this forum post. the program is on Kristen Butela's One Drive. Kristen was posting a work around that is not needed, so get the program from her one drive and then closely follow the installation instructions found oh phagesDB. https://phagesdb.org/DNAMaster/ Best, debbie

Link to this post | posted 03 Mar, 2025 19:57

debbie

Shallee, in a perfect world. you can take 1ul of the sample, make some dilutions, and plate for single plaques. If that doesn't work, there are some additional considerations. (and of course you can start with more than 1ul.) Second step is to pick a single plaque and make a new working lysate, being sure to verify (maybe by pcr) that you have the phage you are interested in. debbie Edited 03 Mar, 2025 19:59

Link to this post | posted 26 Feb, 2025 20:50

debbie

Hi Lisa, In the regions where there are forward and reverse genes called, the previous annotators decided the evidence for forward genes i that region were more compelling. Please be sure to make you decisions based on the evidence that you have. Note that gene prediction models are using a 4 nucleotide sequence to find the most abundant patterns in the big open reading frames, then applying that pattern to the whole genome. The math will break down for smaller genes. My guess is that those regions where genes are called simultaneously on both strands are not very GC rich, so the patterns of ATCG are somewhat equivalent, so they are all getting called. Note to see if one of the programs (Glimmer or GeneMark) calls one strand more than the other. So you may see a bias due to the program's algorithm. debbie Edited 26 Feb, 2025 20:53

Link to this post | posted 26 Feb, 2025 20:46
debbie	It is not common but is definitely seen that Glimmer and GeneMark finds a big open reading frame on opposite strands and then calls most genes in that orientation. These genomes cannot have that many genes in their genomes. Caution is required as to where the other supporting data aligns. Look for the ORFs that have known phage function to build your case for where to call genes.

Link to this post | posted 26 Feb, 2025 16:13

debbie

Hi Adam, hmmmm. This gene is in the region of the minor tail proteins. Minor tail proteins have hydrolase and esterase activity in them. So I was wondering if there what the distal tail looked like for cluster FH. I was hoping to see extra fibers on the end of the tail. First, I believe Cluster FH phages look to be myovirus, even though your particle could be interpreted as a podo. I would check more than one particle to be sure. Looking at the genome, that you see a tape measure protein and many minor tail proteins, I think you will find that this is not a podovirus. My HHPRed results of this gene clearly hit a domain in other phage tail genes. Its location makes me want to call it a tail gene, and my point above that minor tail genes can have hydrolase functions makes it easy for me to call this a minor tail protein. debbie

Link to this post | posted 24 Feb, 2025 01:57
debbie	All AS phages found so far have a tail assembly chaperone preceding the tape measure protein with a programmed translational frameshift. The tail assembly chaperone is typically found in the left arm of the genome around gp13-15. It is a -1 frameshift: G_GG.G_GA.A

Link to this post | posted 20 Feb, 2025 17:47
debbie	Hi Nikki, You have done what I would have done. It is, I think, a Windows error. Have you closed and re-opened DNA Master? You may have to re-start Windows. if you send me the file, i can make it for you. debbie

Link to this post | posted 20 Feb, 2025 15:18
debbie	Hi Jennifer, Have you done any EMs of SwissCheezer? Or looked at EMs of phages of this cluster? I can finish the answer, but I would rather you look there first and let me know what you think. Thanks, debbie

Link to this post | posted 19 Feb, 2025 20:03

debbie

Arturo, Hi. i don't think that the lysis cassette is a reflection of virion morphology. Many phages in our collections have 2 gene that do the function(s) of the endolysins. the mycobacteriophage papers on endolysins are helpful to determine the domains of the endolysins. the exact structure of the cell well is not as clearly understood in the Microbacterium, so we have not seen the gene that corresponds to lysin B. But we have seen the lysin A (endolysin) function to be across 2 separate genes, just like this. Call you genes endolysin, but then list the prominent domain that it is hitting. https://pubmed.ncbi.nlm.nih.gov/22470512/ debie

Link to this post | posted 12 Feb, 2025 23:55
debbie	Nope. Are you experimentally determining them? Are they like other published AttPs? It is a good question to ask. For now, it can be kept in the DNA Master files the annotation notes that you submit on phagesDB. debbie