Link to this post | posted 23 Dec, 2021 20:49

debbie

Chris, I added this, but I have a question. Are there 2 endolysin genes in this genome? Lysin genes are notorious for not having all domains match existing domains in the databases that we use. By that I mean, we can call a lysin A because ONE of the domains hit previously studied endolysins. so in the case of the gene you are interested in, is it possible that the 34% not covered is the missing protease domain? so the gene is a whole endolysin and not a gene broken into 2 pieces? I would recommend that we call the gene an endolysin if no protease domain is identified. Does that make sense? Would you also provide the genome/gene number to list it is an example int he function list. Thanks, debbie Edited 23 Dec, 2021 20:50

Link to this post | posted 17 Dec, 2021 17:45
debbie	Hi Ping, We have seen this before. I would call gene 2 "terminase, small subunit", gene 3 "terminase large subunit (ATPase domain)", and gene 4 "terminase large subunit (nuclease domain)". We have looked into this with the cluster AY phages and worked on a case study to show our investigation. It is still in draft form, but may be helpful. It is attached. 789Kb

Link to this post | posted 13 Dec, 2021 18:06
debbie	Mary, Hi. Are you running as administrator? debbie

Link to this post | posted 13 Dec, 2021 17:44
debbie	Hi all, Did you go back through this forum post and unpack and repack your DNA Master file? That is the typical way to fix this. https://seaphagesbioinformatics.helpdocsonline.com/article-104 It is tedious, so be patient. Let me know if that corrects the problem. If not, please send me the file. Thanks, debbie

Link to this post | posted 12 Dec, 2021 00:22
debbie	Maybe the different pham gene is actually in a different frame. What do you think? debbie

Link to this post | posted 03 Dec, 2021 18:48

debbie

Unlike most of the Microbacterium phages we have found to date (12-2-21), Cluster EH phages are temperate and do have a serine integrase. We see 3 different phams of serine integrases in these genomes so far. There are 7 members in this cluster at this time - Floof, GardenState, Gretchen, Honk, IAmGroot, Percival and Zeta1847. Their starts are called correctly. Look for the serine residue ~ 8-20 bases of the start. Serine integrases are sometimes called large recombinases because of their size. A good read is found here: https://pubmed.ncbi.nlm.nih.gov/26350324/

Link to this post | posted 03 Dec, 2021 15:32

debbie

Dear , It is a common morphology for some of our phages to do this. Six rounds of replating a well isolated plaque means you have purified well. Our Cluster A phages are notorious for doing this. I think that they are the same plaque morphology and it is the landscape differences that do this OR just as likely, it is a cell growth issue and not cells are in the same growth period when infection started so some plaques have a head start on the others. What is 'dangerous' is to 'over-purify' and find the clear plaque mutant that has lost /integration function. If there is a genotypic difference between the 2 populations, a good Illumina sequence run may show it. debbie

Link to this post | posted 19 Nov, 2021 17:22
debbie	Thanks Chris!

Link to this post | posted 19 Nov, 2021 16:11

debbie

Lee, Great forum post. Thanks for having a complete set of data. You are correct that the checker message is confusing. Thanks for reviewing. I think that this tRNA is legit and the coordinates are 99206-99276, tRNA-Gln(gat). There is no way around this in the checker and I am will contact Christian to push this one through. Sometimes, you may receive additional error message, but just ignore. it is ready to go in. Best, debbie

Link to this post | posted 12 Nov, 2021 14:34

debbie

Hi Amanda, Kyle's answer reflects our experiences too. So while I am not sure the details for how best to get phage yields from any of these hosts, figuring that out is legit. The caveat, I think, would be if any are 'ready' for use in the student lab. For us the experience is that we don't have enough experience with any of them to know what best practice is. Denise Monti has used Corynebacterium in the classroom, so you might want to ask her. Good Luck! debbie