Link to this post | posted 24 Mar, 2023 21:28

debbie

Hi Adam, My answer is I don't know. This cluster of genomes is not like the others. It is a podo and I am not sure it fits the canonical features of podos. In order to answer this question, a review of podovirus structures is warranted. What are the canonical featured of them? So a deep dive into the literature is warranted here. Next, the cryoEM data is pouring in to the PDB, so what we have access to now vs when this cluster first appeared has grown. Next, in the literature you can find good reviews about podos and myos tails(and learn that the tail structures of siphos is a bit more elusive). We'll need to apply that to this. Next, your are cherry-picking one gene is a very non-canonical place in the genome. Structural genes frequently hit heads and tails and until we can clearly determine what we have going on here, I recommend that we prudently wait on jumping the gun. Finally, the same terminology doesn't always cross from sipho, myo and podo. So some thought will be required. Keep us posted as you do your investigations. debbie

Link to this post | posted 23 Mar, 2023 15:58
debbie	Veronique, I believe that this issue is outside of DNA Master's reach. Something it wanted to contact couldn't. (or something like that.) Regardless, I would restart my computer and then bet my nickel that it will disappear. debbie

Link to this post | posted 19 Mar, 2023 23:05
debbie	Using the phamerator numbers I would likely not call either 39 or 40. 34296 - 34397 has enough (though slight) coding potential for me to consider calling this one. No Other FF has this sequence. Note that this genome has 2 tRNAs, make sure there is not any overlap between genes/tRNAs.

Link to this post | posted 18 Mar, 2023 00:05
debbie	Excellent! Yay!

Link to this post | posted 17 Mar, 2023 23:59
debbie	Hi! I could call 41673-41810. There is a nib of coding potential and that if fits with 4bp overlaps at both ends is compelling. I think the coding potential is there for 41974-42225. However, I am not sure about the last one 42293-42394. I don't think that i would call it. debbie

Link to this post | posted 17 Mar, 2023 16:23
debbie	Amanda, This sounds like a connection is busted between the windows tools DNA Master uses and DNA Master. You have to get them back. The simplest way forward may be to totally uninstall and then reinstall it on your machine. If you haven't turned your machine off and then back on, that might be a first try. debbie

Link to this post | posted 17 Mar, 2023 14:51

debbie

Nikki, That is not entirely correct. Some folks capture all of the slip in one region, where it is to be slipped between the 2 (meaning the annotation of the shared nucleotide occurs in the middle). Because the slippage happens around a lot of glycines, the sequence can look right but be actually annotated incorrectly. In this particular case, there are 2 predicted overlapping slippery sequences. The run of Gs and the canonical XXXYYYZ. In this particular case, I can't tell which one is used without some bench work. debbie Edited 17 Mar, 2023 16:07

Link to this post | posted 17 Mar, 2023 01:00
debbie	Nikki, Hi. Both choices produce the same protein sequence and unless the experiment is done to label the amino acids in the sequence, I don't think one is better than the other. There is no GN or GK decision, it is whether the second G in the final sequence of AGGK is in the first frame or the second. Does that make sense? debbie

Link to this post | posted 16 Mar, 2023 23:42

debbie

Ellen, Hi. There is no data that supports "secreted protein". I am not sure where that came from, but is no longer in use. when you run this protein through DeepTMHMM, it does have a single transmembrane domain, so "membrane protein" can be assigned to this protein. Note that when you look at pham data on phagesDB, you don't find "secreted protein" as a function call across the pham but the NCBI blast hits that do call it are from RefSeqs. So it appears that the RefSeq's at NCBI are not as current. Supporting data to a call will be present outside of blast if it is a true call (most of the time)> debbie

Link to this post | posted 14 Mar, 2023 19:33

debbie

Hi Ellen, I have attached a screen shot of my HHPRed. The significant hits are 1. e-27/ probability score of 99.9 to gp15 of D29 (sometimes there are hits to gp# that do not connect to a phage, but in this case, hitting the prototypic phage D29, I would recommend paying attention to it). gp15 of D29 is its capsid maturation protease (CMP). Additionally, i can be skeptical of any minor capsid hits, because many times it is hitting some component that i know is structural but can't tell if it is heads or tails. BUT, having said that, hitting minor capsid doesn't negate it being the CMP. Any hits to MuF are problematic. It does not appear to be structural (can't be found in come cryo EM that Simon White has done) and appears to always be part of something else. But never really hits the something else. But maybe MuF is Mu's version of the capsid assembly protein? Is there an instance of both MuF and CMP are present? I would easily call this gene capsid maturation protease. debbie