Soft clipped reads not displaying properly.

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Soft clipped reads not displaying properly.

Faye Rodgers

Hi Apollo developers,

 

I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.

 

So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.

Let me know if you need any further details.

 

Many thanks!

 

Faye

 

-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE.



This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
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Soft_clipped.png (109K) Download Attachment
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Re: Soft clipped reads not displaying properly.

nathandunn

Faye,

We’ll try to isolate this stuff on our end with the data you provided.  

Some things to try (so that we can isolate the issues ahead of time), if possible:

- Can you share your trackList.json object?
- Try to remove the NeatFeatures plugins.  
- Change the track type to Alignments2 (uses canvas)
- Have you tried this with straight JBrowse (trying to isolate the issue).  It should work with the same data directories. 

I’ve created the issue to track progress on this:


Nathan


On Oct 13, 2017, at 8:30 AM, Faye Rodgers <[hidden email]> wrote:

Hi Apollo developers,
 
I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.
 
So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.
Let me know if you need any further details.
 
Many thanks!
 
Faye
 
-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <Soft_clipped.png>


This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 






This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.

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Re: Soft clipped reads not displaying properly.

Deepak Unni
Hi Faye,

The problem here is two fold:

1. NeatHTMLFeatures and NeatCanvasFeatures will lead to incorrect rendering of BAM reads. Since both of these track types are still experimental, I would recommend removing these plugins.
2. There is a bug in DraggableAlignments track type in Apollo. If you change your track configuration from WebApollo/View/Track/DraggableAlignments to JBrowse/View/Track/Alignments2 (as Nathan suggested), you should be able to see the long reads get rendered properly.

I have created an issue for addressing the bug with DraggableAlignments - https://github.com/GMOD/Apollo/issues/1788

Cheers,

Deepak

On Fri, Oct 13, 2017 at 12:39 PM, Nathan Dunn <[hidden email]> wrote:

Faye,

We’ll try to isolate this stuff on our end with the data you provided.  

Some things to try (so that we can isolate the issues ahead of time), if possible:

- Can you share your trackList.json object?
- Try to remove the NeatFeatures plugins.  
- Change the track type to Alignments2 (uses canvas)
- Have you tried this with straight JBrowse (trying to isolate the issue).  It should work with the same data directories. 

I’ve created the issue to track progress on this:


Nathan


On Oct 13, 2017, at 8:30 AM, Faye Rodgers <[hidden email]> wrote:

Hi Apollo developers,
 
I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.
 
So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.
Let me know if you need any further details.
 
Many thanks!
 
Faye
 
-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <Soft_clipped.png>


This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 






This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.





--
Research Analyst
S104A Animal Science Research Center,
University of Missouri, Columbia




This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.

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Re: Soft clipped reads not displaying properly.

Faye Rodgers

Hi Nathan, Hi Deepak,

 

Thanks for the tips. We haven’t quite solved the problem.

 

We have:

-Removed the NeatHTMLFeatures and NeatCanvasFeatures plugins

-Checked the track type in trackList.json: it was actually already set to JBrowse/View/Track/Alignments2. I will share the trackList.json object with you separately.

 

We also updated to the latest version.

 

See the attached image: it looks like the outline of the read gets drawn now, but the aligned bits don’t.

 

Cheers,

Faye

 

 

 

From: <[hidden email]> on behalf of Deepak Unni <[hidden email]>
Reply-To: "[hidden email]" <[hidden email]>
Date: Saturday, October 14, 2017 at 16:54
To: "[hidden email]" <[hidden email]>
Cc: Alan Tracey <[hidden email]>
Subject: Re: [apollo] Soft clipped reads not displaying properly.

 

Hi Faye,

 

The problem here is two fold:

 

1. NeatHTMLFeatures and NeatCanvasFeatures will lead to incorrect rendering of BAM reads. Since both of these track types are still experimental, I would recommend removing these plugins.

2. There is a bug in DraggableAlignments track type in Apollo. If you change your track configuration from WebApollo/View/Track/DraggableAlignments to JBrowse/View/Track/Alignments2 (as Nathan suggested), you should be able to see the long reads get rendered properly.

 

I have created an issue for addressing the bug with DraggableAlignments - https://github.com/GMOD/Apollo/issues/1788

 

Cheers,

 

Deepak

 

On Fri, Oct 13, 2017 at 12:39 PM, Nathan Dunn <[hidden email]> wrote:

 

Faye,

 

We’ll try to isolate this stuff on our end with the data you provided.  

 

Some things to try (so that we can isolate the issues ahead of time), if possible:

 

- Can you share your trackList.json object?

- Try to remove the NeatFeatures plugins.  

- Change the track type to Alignments2 (uses canvas)

- Have you tried this with straight JBrowse (trying to isolate the issue).  It should work with the same data directories. 

 

I’ve created the issue to track progress on this:

 

 

Nathan

 



On Oct 13, 2017, at 8:30 AM, Faye Rodgers <[hidden email]> wrote:

 

Hi Apollo developers,

 

I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.

 

So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.

Let me know if you need any further details.

 

Many thanks!

 

Faye

 

-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <Soft_clipped.png>


This list is for the Apollo Annotation Editing Tool. Info at 
http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to 
[hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 

 





This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.



 

--

Research Analyst
S104A Animal Science Research Center,
University of Missouri, Columbia

-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE.



This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.


soft_clipped_2.png (95K) Download Attachment
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Re: Soft clipped reads not displaying properly.

Deepak Unni
Hi Faye,

Oh, you are referring to the annotation created from the soft-clipped BAM reads.
Thanks for the image. I'll take a look and see what is going on.

Cheers,

Deepak

On Mon, Oct 16, 2017 at 6:06 AM, Faye Rodgers <[hidden email]> wrote:

Hi Nathan, Hi Deepak,

 

Thanks for the tips. We haven’t quite solved the problem.

 

We have:

-Removed the NeatHTMLFeatures and NeatCanvasFeatures plugins

-Checked the track type in trackList.json: it was actually already set to JBrowse/View/Track/Alignments2. I will share the trackList.json object with you separately.

 

We also updated to the latest version.

 

See the attached image: it looks like the outline of the read gets drawn now, but the aligned bits don’t.

 

Cheers,

Faye

 

 

 

From: <[hidden email]> on behalf of Deepak Unni <[hidden email]>
Reply-To: "[hidden email]" <[hidden email]>
Date: Saturday, October 14, 2017 at 16:54
To: "[hidden email]" <[hidden email]>
Cc: Alan Tracey <[hidden email]>
Subject: Re: [apollo] Soft clipped reads not displaying properly.

 

Hi Faye,

 

The problem here is two fold:

 

1. NeatHTMLFeatures and NeatCanvasFeatures will lead to incorrect rendering of BAM reads. Since both of these track types are still experimental, I would recommend removing these plugins.

2. There is a bug in DraggableAlignments track type in Apollo. If you change your track configuration from WebApollo/View/Track/DraggableAlignments to JBrowse/View/Track/Alignments2 (as Nathan suggested), you should be able to see the long reads get rendered properly.

 

I have created an issue for addressing the bug with DraggableAlignments - https://github.com/GMOD/Apollo/issues/1788

 

Cheers,

 

Deepak

 

On Fri, Oct 13, 2017 at 12:39 PM, Nathan Dunn <[hidden email]> wrote:

 

Faye,

 

We’ll try to isolate this stuff on our end with the data you provided.  

 

Some things to try (so that we can isolate the issues ahead of time), if possible:

 

- Can you share your trackList.json object?

- Try to remove the NeatFeatures plugins.  

- Change the track type to Alignments2 (uses canvas)

- Have you tried this with straight JBrowse (trying to isolate the issue).  It should work with the same data directories. 

 

I’ve created the issue to track progress on this:

 

 

Nathan

 



On Oct 13, 2017, at 8:30 AM, Faye Rodgers <[hidden email]> wrote:

 

Hi Apollo developers,

 

I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.

 

So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.

Let me know if you need any further details.

 

Many thanks!

 

Faye

 

-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <Soft_clipped.png>


This list is for the Apollo Annotation Editing Tool. Info at 
http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to 
[hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 

 





This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.



 

--

Research Analyst
S104A Animal Science Research Center,
University of Missouri, Columbia

-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE.




This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.





--
Research Analyst
S104A Animal Science Research Center,
University of Missouri, Columbia




This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.

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Re: Soft clipped reads not displaying properly.

nathandunn
In reply to this post by Faye Rodgers

Just to follow-up for folks on the list who are interested in how to align the PacBio reads.

The solution was to clean up the BAM files and remove CIGAR strings and sequences.

Another suggestion from the Elsik Lab was to produce a GFF from the reads to use during annotation.

Nathan


On Oct 16, 2017, at 4:06 AM, Faye Rodgers <[hidden email]> wrote:

Hi Nathan, Hi Deepak,
 
Thanks for the tips. We haven’t quite solved the problem.
 
We have:
-Removed the NeatHTMLFeatures and NeatCanvasFeatures plugins
-Checked the track type in trackList.json: it was actually already set to JBrowse/View/Track/Alignments2. I will share the trackList.json object with you separately.
 
We also updated to the latest version.
 
See the attached image: it looks like the outline of the read gets drawn now, but the aligned bits don’t.
 
Cheers,
Faye
 
 
 
From: <[hidden email]> on behalf of Deepak Unni <[hidden email]>
Reply-To: "[hidden email]" <[hidden email]>
Date: Saturday, October 14, 2017 at 16:54
To: "[hidden email]" <[hidden email]>
Cc: Alan Tracey <[hidden email]>
Subject: Re: [apollo] Soft clipped reads not displaying properly.
 
Hi Faye,
 
The problem here is two fold:
 
1. NeatHTMLFeatures and NeatCanvasFeatures will lead to incorrect rendering of BAM reads. Since both of these track types are still experimental, I would recommend removing these plugins.
2. There is a bug in DraggableAlignments track type in Apollo. If you change your track configuration from WebApollo/View/Track/DraggableAlignments to JBrowse/View/Track/Alignments2 (as Nathan suggested), you should be able to see the long reads get rendered properly.
 
I have created an issue for addressing the bug with DraggableAlignments - https://github.com/GMOD/Apollo/issues/1788
 
Cheers,
 
Deepak
 
On Fri, Oct 13, 2017 at 12:39 PM, Nathan Dunn <[hidden email]> wrote:
 
Faye,
 
We’ll try to isolate this stuff on our end with the data you provided.  
 
Some things to try (so that we can isolate the issues ahead of time), if possible:
 
- Can you share your trackList.json object?
- Try to remove the NeatFeatures plugins.  
- Change the track type to Alignments2 (uses canvas)
- Have you tried this with straight JBrowse (trying to isolate the issue).  It should work with the same data directories. 
 
I’ve created the issue to track progress on this:
 
 
Nathan
 


On Oct 13, 2017, at 8:30 AM, Faye Rodgers <[hidden email]> wrote:
 
Hi Apollo developers,
 
I'm working on the whipworm school project for WormBase. We've discovered a bug in the way that Apollo displays some full transcript RNASeq reads. The attached image should illustrate the problem. We think we've narrowed the issue down to reads that are soft-clipped at the 5' end: the alignments themselves don't get drawn, only the errors. I will send you a link to the BAM file separately.
 
So far we've been dealing with this by filtering out clipped reads. That's not really a long term solution as we have a lot of trans-splicing (and hence clipped reads) in nematodes.
Let me know if you need any further details.
 
Many thanks!
 
Faye
 
-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <Soft_clipped.png>


This list is for the Apollo Annotation Editing Tool. Info at 
http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to 
[hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 

 





This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.



 
-- 
Research Analyst
S104A Animal Science Research Center,
University of Missouri, Columbia
-- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE. <soft_clipped_2.png>


This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank. 






This list is for the Apollo Annotation Editing Tool. Info at http://genomearchitect.org/
If you wish to unsubscribe from the Apollo List: 1. From the address with which you subscribed to the list, send a message to [hidden email] | 2. In the subject line of your email type: unsubscribe apollo | 3. Leave the message body blank.