SEA-PHAGES | All posts created by cdshaffer

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Link to this post \| posted 17 Mar, 2020 18:37
cdshaffer	I just searched the phamerator database. There are no genes from amy phage with host streptomyces that have lysin B as an annotation.

Posted in: Cluster BD Annotation Tips → lysin A

Link to this post \| posted 10 Mar, 2020 16:41
cdshaffer	There is a lot of help on the "data" page at phagesdb: Data page If you or someone you know has basic scripting skills you can use the API at phagesdb. It gives you the most flexibility and reproducibility if you want to repeat the experiment later. Failing that I would use the bulk system at NCBI. Here is a shorthand protocol I would use: 1. Get "Full tab delimited all phages spreadsheet" from the data page linked above. 2. Use your or a friend's excel skills to filter on cluster/subcluster to create a list with just the phage you want and select and copy the accession numbers column to clipboard 3. Go to Genbank and search the "nucleotide" database with the complete list of all the accession numbers (just paste the whole list in the little search box) 4. On the results page use the tiny menus near the top to change "Summary" to FASTA and then use the "Send to" menu and select "File". You should get a concatenated fasta file wherever your browser downloads. File will be named "sequence.fasta" which you should rename to something move appropriate like "A1_phages.fasta"

Link to this post | posted 10 Mar, 2020 16:41

cdshaffer

There is a lot of help on the "data" page at phagesdb: Data page

If you or someone you know has basic scripting skills you can use the API at phagesdb. It gives you the most flexibility and reproducibility if you want to repeat the experiment later.

Failing that I would use the bulk system at NCBI. Here is a shorthand protocol I would use:

1. Get "Full tab delimited all phages spreadsheet" from the data page linked above.
2. Use your or a friend's excel skills to filter on cluster/subcluster to create a list with just the phage you want and select and copy the accession numbers column to clipboard
3. Go to Genbank and search the "nucleotide" database with the complete list of all the accession numbers (just paste the whole list in the little search box)
4. On the results page use the tiny menus near the top to change "Summary" to FASTA and then use the "Send to" menu and select "File". You should get a concatenated fasta file wherever your browser downloads. File will be named "sequence.fasta" which you should rename to something move appropriate like "A1_phages.fasta"

Posted in: Phage Biology → Downloading cluster specific fasta files

Link to this post \| posted 09 Mar, 2020 16:28
cdshaffer	OK the rule that "you must have a B, in order to have an A" makes sense to me. Thanks for the clarification.

Posted in: Cluster BD Annotation Tips → lysin A

Link to this post \| posted 09 Mar, 2020 00:38
cdshaffer	Based on this posting from Feb 2020: https://seaphages.org/forums/topic/5000/ Which I will repost here: `Just looked at 2 cluster AO2 phages, JKerns and StevieBAY. StevieBAY's gene 28 is an endolysin. It possesses all of domains we expect to see in an endolysin. Gene 64 codes for the CHAP endopeptidase domain of LysK (a Staphylococcus phage lysin gene). It is reported out as an endolysin also. We won't call either of them lysin A (Reserved only for mycobacteriophages for now, unless the phage genomes contains a lysin A and a lysin B.` And this in the approved terms (as of Feb 2020): `some arthrobacter and streptomyces phages have a single endolysin with domains not found in the Mycobacteriophages (like the CHAP domain). use "endolysin" rather than lysin A if the phage does not infect Mycobacterium and no lysin b can be identified.` The BD6 phage appear to have a lysin which is a very high quality HHpred hit (99.8 Prob, 97.2 coverage) to the N-ACETYLMURAMOYL-L-ALANINE AMIDASE crystal (2WKX_A) should annotation be the approved term "endolysin" or "lysin A, N-acetylmuramoyl-L-alanine amidase domain" Edited 09 Mar, 2020 00:50

Link to this post | posted 09 Mar, 2020 00:38

cdshaffer

Based on this posting from Feb 2020:
https://seaphages.org/forums/topic/5000/

Which I will repost here:

Just looked at 2 cluster AO2 phages, JKerns and StevieBAY. StevieBAY's
gene 28 is an endolysin. It possesses all of domains we expect to see in an
endolysin. Gene 64 codes for the CHAP endopeptidase domain of LysK (a
Staphylococcus phage lysin gene). It is reported out as an endolysin also.
We won't call either of them lysin A (Reserved only for mycobacteriophages
for now, unless the phage genomes contains a lysin A and a lysin B.

And this in the approved terms (as of Feb 2020):

some arthrobacter and streptomyces phages have a single endolysin
with domains not found in the Mycobacteriophages (like the CHAP domain).
use "endolysin" rather than lysin A if the phage does not infect
Mycobacterium and no lysin b can be identified.

The BD6 phage appear to have a lysin which is a very high quality HHpred hit (99.8 Prob, 97.2 coverage) to the N-ACETYLMURAMOYL-L-ALANINE AMIDASE crystal (2WKX_A)

should annotation be the approved term "endolysin" or "lysin A, N-acetylmuramoyl-L-alanine amidase domain"

Edited 09 Mar, 2020 00:50

Posted in: Cluster BD Annotation Tips → lysin A

Link to this post \| posted 12 Feb, 2020 23:30
cdshaffer	Just a reminder to everyone that the most recent updates to Starterator were designed to improve the whole phage reports. These whole phage reports steamline some of the output that is needed for a whole pham report but is irrelevant if all you care about is annotating one particular phage. Unfortunately I cannot pre-compute whole starterator reports, its just too computationally intensive, but I am happy to generate a whole phage report for anyone by request. Simply post a follow-up message to this post or send me an email. All I need is the name of the phage and I can generate a report based on the initial auto-annotation. Be aware that for phage in large clusters it can take several hours of computation so expect some delay between request and response but it only takes me about 45 seconds to start the run and another 75 seconds to post the results so don't feel like this is a huge ask, happy to help. Also, you may find it helpful to run a whole phage report on a phage after you have worked on annotation and want to rerun the whole phage report based on those newer annotations. I can do that easily as well. Simply email me either your DNA Master file, or if you are using PECAAN export and send me the "CDS function" report and I can run a whole phage starterator report based on that new annotation set. Edited 13 Feb, 2020 03:26

Link to this post | posted 12 Feb, 2020 23:30

cdshaffer

Just a reminder to everyone that the most recent updates to Starterator were designed to improve the whole phage reports. These whole phage reports steamline some of the output that is needed for a whole pham report but is irrelevant if all you care about is annotating one particular phage. Unfortunately I cannot pre-compute whole starterator reports, its just too computationally intensive, but I am happy to generate a whole phage report for anyone by request. Simply post a follow-up message to this post or send me an email. All I need is the name of the phage and I can generate a report based on the initial auto-annotation. Be aware that for phage in large clusters it can take several hours of computation so expect some delay between request and response but it only takes me about 45 seconds to start the run and another 75 seconds to post the results so don't feel like this is a huge ask, happy to help.

Also, you may find it helpful to run a whole phage report on a phage after you have worked on annotation and want to rerun the whole phage report based on those newer annotations. I can do that easily as well. Simply email me either your DNA Master file, or if you are using PECAAN export and send me the "CDS function" report and I can run a whole phage starterator report based on that new annotation set.

Edited 13 Feb, 2020 03:26

Posted in: Starterator → whole phage starterator reports

Link to this post \| posted 08 Feb, 2020 00:29
cdshaffer	Yes there was a new version of the database posted this morning. Since each site updates as it can things can get out of sync. Not sure how you generated what you are calling your "gene list" but pecaan and phagesdb were updated earlier today so the links out of pecaan and out of phagesdb were pointing to pages that did not exist until starterator was also updated to the new version 339. This is how you can get the info on which version each site is on (links will only work between two sites if they are on the same version): phagesdb see: http://phamerator.webfactional.com/databases_Hatfull/Actino_Draft.version starterator see: http://phages.wustl.edu/starterator/database.version pecaan: look on any "pham maps" page just above the map phamerator.org: open the pull down menu in the top left Edited 08 Feb, 2020 00:30

Link to this post | posted 08 Feb, 2020 00:29

cdshaffer

Yes there was a new version of the database posted this morning. Since each site updates as it can things can get out of sync.
Not sure how you generated what you are calling your "gene list" but pecaan and phagesdb were updated earlier today so the links out of pecaan and out of phagesdb were pointing to pages that did not exist until starterator was also updated to the new version 339. This is how you can get the info on which version each site is on (links will only work between two sites if they are on the same version):
phagesdb see: http://phamerator.webfactional.com/databases_Hatfull/Actino_Draft.version
starterator see: http://phages.wustl.edu/starterator/database.version
pecaan: look on any "pham maps" page just above the map
phamerator.org: open the pull down menu in the top left

Edited 08 Feb, 2020 00:30

Posted in: Starterator → Pham not found in Starterator

Link to this post \| posted 05 Feb, 2020 20:13
cdshaffer	I am adding an online link so it is much easier to see the current Starterator database version. This link should always give the most recent database version that has been published: http://phages.wustl.edu/starterator/database.version

Posted in: Starterator → Database Version for starterator

Link to this post \| posted 05 Feb, 2020 16:20
cdshaffer	Yes a typo of any kind can keep the system from changing to a green check. One simple error that is hard to detect is if there is a space character to the beginning or end of the function. The other thing worth mentioning but cannot explain your high rate is that the approved function list changes quite quickly but Pecaan only updates its list periodically. So again you can have synchronization issues. It is not uncommon for us to have an X or two by the end of an annotation because of this. Not sure why you are getting up to 1/3rd. So my suggestion is the same as JoAnn, encourage the students to type just a few characters in the function and then select the correct entry from the dropdown. Edited 05 Feb, 2020 16:26

Link to this post | posted 05 Feb, 2020 16:20

cdshaffer

Yes a typo of any kind can keep the system from changing to a green check. One simple error that is hard to detect is if there is a space character to the beginning or end of the function. The other thing worth mentioning but cannot explain your high rate is that the approved function list changes quite quickly but Pecaan only updates its list periodically. So again you can have synchronization issues. It is not uncommon for us to have an X or two by the end of an annotation because of this. Not sure why you are getting up to 1/3rd. So my suggestion is the same as JoAnn, encourage the students to type just a few characters in the function and then select the correct entry from the dropdown.

Edited 05 Feb, 2020 16:26

Posted in: PECAAN → PECAAN Down?

Link to this post \| posted 03 Feb, 2020 03:41
cdshaffer	This is all happening because of database synchronization issues. Right now pecaan is on 336 and the rest of us have already updated to version 337. You can tell which version in various ways. Since phagesdb.org is always in sync with the web server that publishes the database, you can bookmark this page to see the most current version of the database: http://phamerator.webfactional.com/databases_Hatfull/Actinobacteriophage.version For phamerator.org there is a pulldown menu in the top right with the database version number; for Pecaan the version is noted just above the map on the Pham Maps tab; for starterator the version is noted just after the run date near the top of the text section of the report so any working report can be used. You can also just go to the URL "phages.wustl.edu/<version>/" to see if that version has been posted. For example, to check if version 337 is available go here: http://phages.wustl.edu/337/ All those reports are specific for version 337. Since Pecaan is on an older version than starterator there is a work around. Just change the "starterator" in the url to the database version number. So when pecaan is using the old 336 and it tries to use an out of date link, this one for example http://phages.wustl.edu/starterator/Pham99984Report.pdf it fails with a 404, so knowing Pecaan is using version 336 replace the "starterator" in the URL with the version number like this: http://phages.wustl.edu/336/Pham99984Report.pdf.pdf and now it works. When starterator is behind pecaan there is not much that can be done but wait until I post the newest set; but please feel free to send me an email and I can at least give you an ETA. P.S. the above links may not work correctly once version 338 of the database is released. Edited 03 Feb, 2020 21:28

Link to this post | posted 03 Feb, 2020 03:41

cdshaffer

This is all happening because of database synchronization issues. Right now pecaan is on 336 and the rest of us have already updated to version 337. You can tell which version in various ways.

Since phagesdb.org is always in sync with the web server that publishes the database, you can bookmark this page to see the most current version of the database:

http://phamerator.webfactional.com/databases_Hatfull/Actinobacteriophage.version

For phamerator.org there is a pulldown menu in the top right with the database version number; for Pecaan the version is noted just above the map on the Pham Maps tab; for starterator the version is noted just after the run date near the top of the text section of the report so any working report can be used. You can also just go to the URL "phages.wustl.edu/<version>/" to see if that version has been posted. For example, to check if version 337 is available go here:
http://phages.wustl.edu/337/
All those reports are specific for version 337.

Since Pecaan is on an older version than starterator there is a work around. Just change the "starterator" in the url to the database version number. So when pecaan is using the old 336 and it tries to use an out of date link, this one for example

http://phages.wustl.edu/starterator/Pham99984Report.pdf

it fails with a 404, so knowing Pecaan is using version 336 replace the "starterator" in the URL with the version number like this:
http://phages.wustl.edu/336/Pham99984Report.pdf.pdf

and now it works.

When starterator is behind pecaan there is not much that can be done but wait until I post the newest set; but please feel free to send me an email and I can at least give you an ETA.
P.S. the above links may not work correctly once version 338 of the database is released.

Edited 03 Feb, 2020 21:28

Posted in: PECAAN → PECAAN Down?

Link to this post \| posted 29 Jan, 2020 22:27
cdshaffer	To adjust the VM desktop image use the virtual box View -> Virtual Screen -> Scale to… menu item. When I am on a high res screen like my macbook pro screen I set "Scale to 200%" and when I am on an external monitor I set "Scale to 100%" but feel free to pick whatever scale works best for you.

Posted in: Using WINE to run DNA Master on a Mac → Help with WINE

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