SEA-PHAGES | All posts created by amaya.garciacostas

Link to this post \| posted 03 Jun, 2023 15:21
amaya.garciacostas	Many AZ phage have a small gene (about 120 bp) right before DNA polymerase that is often not called by Glimmer, but can be seen through the coding potential: great case study for students about missed ORFs during the autoannotation! (See Adumb2043 coordinates: 28674 to 28796 for an example)

Posted in: Cluster AZ Annotation Tips → Small gene in gap

Link to this post \| posted 07 Oct, 2022 17:58
amaya.garciacostas	Hi! It looks like "putative thiamine binding protein" is no longer in the functions list, am I correct? We have quite a few genomes in the AZ cluster with that annotation and I am working on correcting them, thanks!

Posted in: Request a new function on the SEA-PHAGES official list → thiamine-binding protein

Link to this post \| posted 07 Oct, 2022 17:58
amaya.garciacostas	Hi! It looks like "putative thiamine binding protein" is no longer in the functions list, am I correct? We have quite a few genomes in the AZ cluster with that annotation and I am working on correcting them, thanks!

Posted in: Request a new function on the SEA-PHAGES official list → thiamine-binding protein

Link to this post \| posted 07 Oct, 2022 17:58
amaya.garciacostas	Hi! It looks like "putative thiamine binding protein" is no longer in the functions list, am I correct? We have quite a few genomes in the AZ cluster with that annotation and I am working on correcting them, thanks!

Posted in: Request a new function on the SEA-PHAGES official list → thiamine-binding protein

Link to this post \| posted 21 Apr, 2022 21:53
amaya.garciacostas	Gene 39 in Aoka also appears to be a DpdA-like tRNA-guanine transglycosylase. BLAST search called it a queuine tRNA ribosyltransferase but of course we didn't see that in the functions list. All of the members of the Pham, for what it is worth, have called it a DpdA-like tRNA-guanine transglycosylase. We tried to do an alignment with Orion20 from the case study, but there is no significant alignment so not great. HHPred also has hits with high coverage and probability and low e value to a Queuine tRNA-ribosyltransferase, so we are ready to call it a DpdA-like tRNA-guanine transglycosylase and just wanted to share this since it seems like you are actively working on this. No gene context is helpful as this gene is surrounded by orphams.

Link to this post | posted 21 Apr, 2022 21:53

amaya.garciacostas

Gene 39 in Aoka also appears to be a DpdA-like tRNA-guanine transglycosylase. BLAST search called it a queuine tRNA ribosyltransferase but of course we didn't see that in the functions list. All of the members of the Pham, for what it is worth, have called it a DpdA-like tRNA-guanine transglycosylase. We tried to do an alignment with Orion20 from the case study, but there is no significant alignment so not great. HHPred also has hits with high coverage and probability and low e value to a Queuine tRNA-ribosyltransferase, so we are ready to call it a DpdA-like tRNA-guanine transglycosylase and just wanted to share this since it seems like you are actively working on this. No gene context is helpful as this gene is surrounded by orphams.

Posted in: Request a new function on the SEA-PHAGES official list → queuine tRNA ribosyltransferase

Link to this post \| posted 05 Apr, 2022 21:11
amaya.garciacostas	We found a gene labeled "portal and MuF-like fusion protein" that is not in the function list. There is only a portal protein on that list but it is not specified what these portal and MuF-like fusion proteins should be called. It is called the portal and MuF-like fusion protein in the Phage Maja and is coming up in NCBI exactly like that for two different hits. What should these be annotated as?

Posted in: Cluster AK Annotation Tips → Portal and MuF-like

Link to this post \| posted 17 Mar, 2022 20:43
amaya.garciacostas	Thanks for this awesome feedback!! I shared it with the students that were working on those genes an it was very helpful. Our class is small so they appreciated having a broader discussion with somebody outside of our small circle. Yay! cdshaffer First, I would say that having an orpham or two in a phage is not to unusual to really worry me. In looking at the phamerator map of all AZ phage I can see that KeAlii has at least 3 orphams, and there are several phage with 1 or 2 orphams so having two orphams is not so unusual as to cause real questions in these genes. The longer gene 54 has such good coding potential I would always call that one. 53 is just long enough to call. See rule 8 guiding Principles.. I would say 35 amino acids is in the grey zone, meaining it requires some evidence other than just an open reading frame to call the gene. However both have pretty good coding potential so I would call both. No BLAST hits is also not so surprising as we already know it is an orpham (i.e. unique among all 400K proteins in phagesd). So while a good BLASTp metch might make you feel more confident there really is a gene here, the lack of a BLAST hit is not good evidence that this region is not a gene. Said more formally, a positive result in a BLASTp is good evidence, a negative result is not good evidence there is no gene, it is simply that BLAST has nothing to say one way or the other. As for overlap, we would call this a 4 base overlap (gap score of -4). Since gene coordinates describe intervals not counts you cannot just subtract the coordinates, you have to adjust by 1. I run across this issue all the time with my students and I have them draw out a tiny sequence with a few "genes" to see the difference between interval math and normal math. Finally, for gene calls, it is better to have a false positive (i.e. call a gene which really isn't there) than it is to have a false negative (miss a gene). So even if I was not sure of gene 53 I would still call it given this rule. So for all the above, I would keep both these genes in the annotation and just be amazed at how diverse the gene collection is among all these phage. I am sure Deb could quote you a few papers that discuss the ideas of phage as "engines of gene creation" and I think, at least for 54, that we could have an example of that.

Link to this post | posted 17 Mar, 2022 20:43

amaya.garciacostas

Thanks for this awesome feedback!! I shared it with the students that were working on those genes an it was very helpful. Our class is small so they appreciated having a broader discussion with somebody outside of our small circle. Yay!

cdshaffer
First, I would say that having an orpham or two in a phage is not to unusual to really worry me. In looking at the phamerator map of all AZ phage I can see that KeAlii has at least 3 orphams, and there are several phage with 1 or 2 orphams so having two orphams is not so unusual as to cause real questions in these genes.

The longer gene 54 has such good coding potential I would always call that one. 53 is just long enough to call. See rule 8 guiding Principles.. I would say 35 amino acids is in the grey zone, meaining it requires some evidence other than just an open reading frame to call the gene. However both have pretty good coding potential so I would call both.

No BLAST hits is also not so surprising as we already know it is an orpham (i.e. unique among all 400K proteins in phagesd). So while a good BLASTp metch might make you feel more confident there really is a gene here, the lack of a BLAST hit is not good evidence that this region is not a gene. Said more formally, a positive result in a BLASTp is good evidence, a negative result is not good evidence there is no gene, it is simply that BLAST has nothing to say one way or the other.

As for overlap, we would call this a 4 base overlap (gap score of -4). Since gene coordinates describe intervals not counts you cannot just subtract the coordinates, you have to adjust by 1. I run across this issue all the time with my students and I have them draw out a tiny sequence with a few "genes" to see the difference between interval math and normal math.

Finally, for gene calls, it is better to have a false positive (i.e. call a gene which really isn't there) than it is to have a false negative (miss a gene). So even if I was not sure of gene 53 I would still call it given this rule.

So for all the above, I would keep both these genes in the annotation and just be amazed at how diverse the gene collection is among all these phage. I am sure Deb could quote you a few papers that discuss the ideas of phage as "engines of gene creation" and I think, at least for 54, that we could have an example of that.

Posted in: Gene or not a Gene → Orpham genes in AZ phage

Link to this post \| posted 10 Mar, 2022 05:18
amaya.garciacostas	My class is annotating VResidence which is an AZ phage closely related to DrSierra. Much to our surprise -given how many AZ phage there are already sequenced-, there are two orpham genes towards the end. Gene 53 (stop at 3711 and gene 54 (stop at 37497). We are inclined to call them genes mostly because of gaps: without them, there is too large of a gap. And they have a 3 bp overlap with the previous gene and with each other … is that chance? We don't think so! But rather than putting this in the notes, we are communicating in real time as requested! Here is the paragraph about it that my student wrote; we investigated the possibility of a different gene being there instead of those two, and the coding potential is a bit noisy: We found for Gene #53 that there seemed to be some coding potential in a different frame sequence. However, in this frame, there was no stop or start that could be identified. When looking at the frames in DNA master, the reverse would overlap too much with Gene #52 to be considered. There were no possible homologs found in Phamerator and there were no hits when locally blasted in the phagesdb. There were no hits in the NCBI nr database as well. For Gene #54, there was a coding potential with a stop and start in the direct sequence. There were no homologs found in Phamerator. The NCBI database also had no hits for this protein sequence. For gene #54, when locally blasted in the phagesbd, all the hits were proteins with an unknown function. HHPred had no hits for either of the sequences. We think there may be a possibility that gene 53 and gene 54 may be the same gene. Attached is an image from the coding potential: Gene 53 is the first gene found in he 3rd frame. Edited 10 Mar, 2022 22:24 120Kb

Link to this post | posted 10 Mar, 2022 05:18

amaya.garciacostas

My class is annotating VResidence which is an AZ phage closely related to DrSierra. Much to our surprise -given how many AZ phage there are already sequenced-, there are two orpham genes towards the end. Gene 53 (stop at 3711 smile

and gene 54 (stop at 37497). We are inclined to call them genes mostly because of gaps: without them, there is too large of a gap. And they have a 3 bp overlap with the previous gene and with each other … is that chance? We don't think so! But rather than putting this in the notes, we are communicating in real time as requested!

Here is the paragraph about it that my student wrote; we investigated the possibility of a different gene being there instead of those two, and the coding potential is a bit noisy:

We found for Gene #53 that there seemed to be some coding potential in a different frame sequence. However, in this frame, there was no stop or start that could be identified. When looking at the frames in DNA master, the reverse would overlap too much with Gene #52 to be considered. There were no possible homologs found in Phamerator and there were no hits when locally blasted in the phagesdb. There were no hits in the NCBI nr database as well. For Gene #54, there was a coding potential with a stop and start in the direct sequence. There were no homologs found in Phamerator. The NCBI database also had no hits for this protein sequence. For gene #54, when locally blasted in the phagesbd, all the hits were proteins with an unknown function. HHPred had no hits for either of the sequences. We think there may be a possibility that gene 53 and gene 54 may be the same gene.

Attached is an image from the coding potential: Gene 53 is the first gene found in he 3rd frame.

Edited 10 Mar, 2022 22:24

Posted in: Gene or not a Gene → Orpham genes in AZ phage

Link to this post \| posted 22 Feb, 2022 00:19
amaya.garciacostas	nancyguild Dan, One of our students is having trouble loading virtual box on his Mac Book Air (M1, 2020) Mac OS Monterey version 12.1 He gets this message: "Unsupported hardware architecture detected! The installer has detected an unsupported architecture. Virtual box only runs on the amd64 architecture”. The information on the web is confusing in terms of the ability to run virtual box on the M1 macs. Some seem to claim a workaround, others say there is no way to run VB on this type of Mac. Do you have any ideas how to help hime? Thanks! Hi Nancy, Not sure if you had this resolved, but you can go to : https://seaphages.org/forums/topic/5256/ where it was discussed that Virtual box does not work in the new Macs. A new software to try is Parallel, though it costs $40 per student. We had the same problem a month ago!

Link to this post | posted 22 Feb, 2022 00:19

amaya.garciacostas

nancyguild
Dan, One of our students is having trouble loading virtual box on his Mac Book Air (M1, 2020) Mac OS Monterey version 12.1 He gets this message: "Unsupported hardware architecture detected! The installer has detected an unsupported architecture. Virtual box only runs on the amd64 architecture”. The information on the web is confusing in terms of the ability to run virtual box on the M1 macs. Some seem to claim a workaround, others say there is no way to run VB on this type of Mac. Do you have any ideas how to help hime?

Thanks!

Hi Nancy,

Not sure if you had this resolved, but you can go to : https://seaphages.org/forums/topic/5256/
where it was discussed that Virtual box does not work in the new Macs. A new software to try is Parallel, though it costs $40 per student. We had the same problem a month ago!

Posted in: DNA Master → DNA Master and Windows 10

Link to this post \| posted 22 Feb, 2022 00:19
amaya.garciacostas	nancyguild Dan, One of our students is having trouble loading virtual box on his Mac Book Air (M1, 2020) Mac OS Monterey version 12.1 He gets this message: "Unsupported hardware architecture detected! The installer has detected an unsupported architecture. Virtual box only runs on the amd64 architecture”. The information on the web is confusing in terms of the ability to run virtual box on the M1 macs. Some seem to claim a workaround, others say there is no way to run VB on this type of Mac. Do you have any ideas how to help hime? Thanks! Hi Nancy, Not sure if you had this resolved, but you can go to : https://seaphages.org/forums/topic/5256/ where it was discussed that Virtual box does not work in the new Macs. A new software to try is Parallel, though it costs $40 per student. We had the same problem a month ago!

Link to this post | posted 22 Feb, 2022 00:19

amaya.garciacostas

nancyguild
Dan, One of our students is having trouble loading virtual box on his Mac Book Air (M1, 2020) Mac OS Monterey version 12.1 He gets this message: "Unsupported hardware architecture detected! The installer has detected an unsupported architecture. Virtual box only runs on the amd64 architecture”. The information on the web is confusing in terms of the ability to run virtual box on the M1 macs. Some seem to claim a workaround, others say there is no way to run VB on this type of Mac. Do you have any ideas how to help hime?

Thanks!

Posted in: DNA Master → DNA Master and Windows 10

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