Link to this post | posted 04 Oct, 2022 15:45

debbie

Hi Stephanie, If you are using 7H10 to make your agar, it contains malachite green and the agar should really be green. Once overcooked and autoclaved, it tends to be more tan than pale green. If this is the cause of the green, there is nothing wrong. If the agar is undercooked or autoclaved, it can appear greener, BUT you will also have no growth and/or contamination issues, which are a bigger problem. Does that fit what you are seeing?

Link to this post | posted 13 Sep, 2022 13:22

debbie

Tammy, We have used Corynebacterium vitaeruminis a bit. I believe we used PYCa system for plating. My recollection is that the original strain has a prophage in it, the Corynebacterium vitaeruminis CAG1 strain is cured of its prophage. Denise Monti has the most experience, so I would ask her. (montidenise2@gmail.com) We have not obtained any NRRL Corynebacterium hosts, but they look like they have a few. Good luck! Keep us posted on how you proceed! best, debbie

Link to this post | posted 12 Sep, 2022 17:25

debbie

Hi Alison, Could it be that you have a mutant of "WT" Arthrobacter sp.? So you still have your original stock in the freezer? You will want to use that until you confirm what the orange culture is. (Maybe a 16S to confirm). I believe G. terrae CAG3 is a mutant of G. terrae 3612, and is also a mutant in the gene that produces the color. In order to use it for phagehunting, you will want to confirm what you have. With propagation, does it stay orange? or does it go back to yellow. (Is this a mutable change or an environmental change?) Did you make any changes in the prep materials (a different brand of agar)? You will want to test your previous phages on the the new host to see if this change is results in a different infection pattern. If it is now a different bacteria than you started with, meaning it is an inherited trait, it is a new strain, subspecies and should be labeled as a new thing. Keep us at Pitt in the loop so that you enter your phage information correctly. We can add a new subspecies to the list if this proves to be true. Show us a picture of the pretty orange! debbie

Link to this post | posted 19 Aug, 2022 23:36
debbie	To the best of knowledge, DNA mater will not work on an M1 Mac. debbie

Link to this post | posted 12 Aug, 2022 15:33
debbie	Hi Amanda, I see no clear evidence for calling this one anything but a Hypothetical Protein. (My best explanation for why some members of the pham are called terminase, small subunit is there was a moment in time when we thought we could call by synteny. But I would not recommend that today.) Best, debbie

Link to this post | posted 09 Aug, 2022 14:47
debbie	I guess I am going to weigh in as grateful for this post. We have a record and that will help as we go forward. For now, I am going to suggest that we do not add DPS family protein into the approved list. I would recommend that this protein be called a Hypothetical Protein for now. debbie

Link to this post | posted 05 Aug, 2022 15:45

debbie

Kathleen, I am going to answer that question. It is because once a function is identified, it gets propagated. Doesn't matter who said it, or what the supporting evidence is. For SMART, our focus this year has really been to work through these mis-identifications. So questions like yours will help us all get there. The best way to identify where the calls are difficult is to look at phams with multiple functional calls. We all need to work through the data and learn a little bit of structural biology/chemistry and biochemistry to make better decisions. Not always clear and we all bring a different understanding. And the answer to your question about phams is no, not all members in a pham will have the same function. Simplistically, I can see why we think that they should. But if we understand how a pham is built, then we would know that all things in the pham are not the same. A simple explanation is the one member may not have a 'domain' that another one has, so part of the gene is homologues, but missing a particular functional domain would imply that it has a different function. Maybe! An important component is that context is important. debbie

Link to this post | posted 02 Aug, 2022 01:07
debbie	Veronique, I just blasted one gene with no issues. there have been no changes in DNA Master for a while now. debbie

Link to this post | posted 02 Aug, 2022 01:04
debbie	Yep. You got it! debbie

Link to this post | posted 01 Aug, 2022 16:43

debbie

Hi Nick, Membrane proteins, especially those containing 1 transmembrane proteins, are tricky. Very much is dependent on how good the program used to call it is. We are formulating a plan to improve how this work is done, but it is still in process. In the meantime, here is the status of reporting transmembrane proteins. As it turns out, as Travis revamped the database that informs phamerator, he removed certain products because they were problematic. These include hypothetical, gp, orf, putative, and many more. Membrane protein was included on the list because of the 'iffy' nature of how membrane proteins are called. So what you describe is exactly correct. and unfortunately, a bit confusing. (Today we would make a different decision about this, but that is what we have for now.) Call membrane proteins to the best of your ability. Hopefully more news to follow soon. Best, debbie