ChIP-seq

[M***@tnw.utwente.nl]

Dear sir / madame,



I've performed Chromatin Immunoprecipitation and had a service company
sequence the fragments with an Illumina Solexa sequencer. After the
sequence reads had passed the quality check, the ELAND program was used
to align the reads to the hg18 provided by UCSC as part of the Illumina
data analysis pipeline. This was also done by the same company.

Thereafter the aligned sequence tags were fed into the FindPeaks
3.1.9.2. package written by the Canada Michael Smith Genome Sciences
Centre in order to 1. analyze the short-read sequences to identify areas
of enrichment of the protein of interest and 2. generating wig files for
use in the UCSC genome browser. The company performed a FindPeaks run
with standard settings. After I adjusted the settings to my own wished,
I performed another FindPeaks run. The wig file that was generated by
the company could be put into the genome browser without any problems.
After uploading the wig file created by myself, it seems at first that
it went all right because the following screen appears:



Manage Custom Tracks







genome: Human assembly: Mar. 2006 [hg18]

Name

Description

Type

Doc



test3_duplicates_subpeaks_len_triangle
<http://genome.ucsc.edu/cgi-bin/hgCustom?hgsid=114649973&hgct_table=ct_t
est3duplicatessubpeakslentriangle>

test3_ht:1.0_FL:triangle_dupe_rds_inc

wiggle_0















But when I want to go to the genome browser, I the following error
message appears:



Internal error (Unmatched ' line 1 of input: missing closing ' ):
removing custom tracks



Do you know by any chance what went wrong? I've already tried the
standard FindPeaks settings the company also used, but it doesn't work
form me.

I hope someone will have an answer for me. Thanks in advance!



Regards,

Marije



Drs. Marije Telgenkamp, MD

PhD candidate Molecular Cell Biology

Department of Science and Technology

University of Twente

Building Zuidhorst, room ZH125

PO box 217

7500 AE Enschede, The Netherlands

Phone: [phone]

Fax: [phone]

***@tnw.utwente.nl



_______________________________________________
Genome maillist - ***@soe.ucsc.edu
http://www.soe.ucsc.edu/mailman/listinfo/genome

[Pauline Fujita]

Hello Marije,

Could you send me the first 8-10 lines of your wig file so that I can
take a look at it? Please send your reply directly to me without cc'ing
the whole list.

Best regards,

Pauline Fujita

UCSC Genome Bioinformatics Group
http://genome.ucsc.edu

Post by M***@tnw.utwente.nl
Dear sir / madame,
I've performed Chromatin Immunoprecipitation and had a service company
sequence the fragments with an Illumina Solexa sequencer. After the
sequence reads had passed the quality check, the ELAND program was used
to align the reads to the hg18 provided by UCSC as part of the Illumina
data analysis pipeline. This was also done by the same company.
Thereafter the aligned sequence tags were fed into the FindPeaks
3.1.9.2. package written by the Canada Michael Smith Genome Sciences
Centre in order to 1. analyze the short-read sequences to identify areas
of enrichment of the protein of interest and 2. generating wig files for
use in the UCSC genome browser. The company performed a FindPeaks run
with standard settings. After I adjusted the settings to my own wished,
I performed another FindPeaks run. The wig file that was generated by
the company could be put into the genome browser without any problems.
After uploading the wig file created by myself, it seems at first that
Manage Custom Tracks
genome: Human assembly: Mar. 2006 [hg18]
Name
Description
Type
Doc
test3_duplicates_subpeaks_len_triangle
<http://genome.ucsc.edu/cgi-bin/hgCustom?hgsid=114649973&hgct_table=ct_t
est3duplicatessubpeakslentriangle>
test3_ht:1.0_FL:triangle_dupe_rds_inc
wiggle_0
But when I want to go to the genome browser, I the following error
removing custom tracks
Do you know by any chance what went wrong? I've already tried the
standard FindPeaks settings the company also used, but it doesn't work
form me.
I hope someone will have an answer for me. Thanks in advance!
Regards,
Marije
Drs. Marije Telgenkamp, MD
PhD candidate Molecular Cell Biology
Department of Science and Technology
University of Twente
Building Zuidhorst, room ZH125
PO box 217
7500 AE Enschede, The Netherlands
Phone: [phone]
Fax: [phone]
_______________________________________________
http://www.soe.ucsc.edu/mailman/listinfo/genome

_______________________________________________
Genome maillist - ***@soe.ucsc.edu
http://www.soe.ucsc.edu/mailman/listinfo/genome

[Anthony Fejes]

Hello Dr. Telgenkamp,

Your posting was forwarded to me by a colleague, and I felt it would be
appropriate to reply to it, despite not being a regular user of this
mailing list, as I currently maintain and develop the FindPeaks software
package.

I have seen the problem you're describing before, and it appears to have
a relatively simple solution - modifying the track line of the wig file
will correct the problem, though it may not be simple for all users
depending on what software you have available to you. In the past, the
solution has been as simple as moving the 'type="wiggle_0"' to follow
immediately after word 'track'.

The problem seems to arise because FindPeaks 3.1.9.2 produces wig files
with the "track" line parameters in a different order from that given in
the wig file documentation,
(http://genome.ucsc.edu/goldenPath/help/wiggle.html). I don't know why
only some files produce the error, through re-ordering the parameters
does seem to resolve the issue for the two examples I've seen.

This problem has been fixed in FindPeaks 3.2, but was not back ported to
FindPeaks 3.1. Unfortunately, FindPeaks 3.2 won't be available as a
recompiled jar file for another week or so.

In any case, if you'd like to send me your wig file (or just the first
10 lines, either should work, if the file isn't too big), I would be
happy to help you find a fix for the problem.

Finally, it was also pointed out to me by my colleague that you seem to
be using a parameter "-minimum 1". You may wish to raise this value for
tracks submitted to the UCSC browser, as this can cause wig files to
become very large, and I understand the UCSC browser may not load wig
files over 300k long. (Peak heights below 2 are rarely informative
anyhow, and significantly add to the size of the file.)

Alternately, you might wish to leave this particular parameter off
entirely, as a minimum peak height of 1 will still provide all peaks
under most circumstances.

In any case, feel free to contact me if you need any further help with
this issue. There is also a FindPeaks mailing list at
vancouvershortr-***@lists.sourceforge.net, which can be used for
any other FindPeaks problems you may encounter in the future.

Cheers,

Anthony Fejes,
BC Genome Sciences Centre

Message: 5
Date: Mon, 27 Oct 2008 17:17:31 +0100
Subject: [Genome] ChIP-seq
Content-Type: text/plain; charset="us-ascii"
Dear sir / madame,
I've performed Chromatin Immunoprecipitation and had a service company
sequence the fragments with an Illumina Solexa sequencer. After the
sequence reads had passed the quality check, the ELAND program was used
to align the reads to the hg18 provided by UCSC as part of the Illumina
data analysis pipeline. This was also done by the same company.
Thereafter the aligned sequence tags were fed into the FindPeaks
3.1.9.2. package written by the Canada Michael Smith Genome Sciences
Centre in order to 1. analyze the short-read sequences to identify areas
of enrichment of the protein of interest and 2. generating wig files for
use in the UCSC genome browser. The company performed a FindPeaks run
with standard settings. After I adjusted the settings to my own wished,
I performed another FindPeaks run. The wig file that was generated by
the company could be put into the genome browser without any problems.
After uploading the wig file created by myself, it seems at first that
Manage Custom Tracks
genome: Human assembly: Mar. 2006 [hg18]
Name
Description
Type
Doc
test3_duplicates_subpeaks_len_triangle
<http://genome.ucsc.edu/cgi-bin/hgCustom?hgsid=114649973&hgct_table=ct_t
est3duplicatessubpeakslentriangle>
test3_ht:1.0_FL:triangle_dupe_rds_inc
wiggle_0
But when I want to go to the genome browser, I the following error
removing custom tracks
Do you know by any chance what went wrong? I've already tried the
standard FindPeaks settings the company also used, but it doesn't work
form me.
I hope someone will have an answer for me. Thanks in advance!
Regards,
Marije
Drs. Marije Telgenkamp, MD
PhD candidate Molecular Cell Biology
Department of Science and Technology
University of Twente
Building Zuidhorst, room ZH125
PO box 217
7500 AE Enschede, The Netherlands
Phone: [phone]
Fax: [phone]

_______________________________________________
Genome maillist - ***@soe.ucsc.edu
http://www.soe.ucsc.edu/mailman/listinfo/genome

[Anthony Fejes]

Just to provide an update to this problem, it seems my initial
suggestion was incorrect. The UCSC browser does not like the header
lines at the top of the wig file. Removing all of the lines before the
one that starts with "track" will correct the problem.

I'm still not certain why this only affects certain files and not
others. It may have to do with the line termination character in the
header, which I'll ensure is not platform dependent in FindPeaks 3.2.

Anthony

Message: 5
Date: Mon, 27 Oct 2008 17:17:31 +0100
Subject: [Genome] ChIP-seq
Content-Type: text/plain; charset="us-ascii"
Dear sir / madame,
I've performed Chromatin Immunoprecipitation and had a service company
sequence the fragments with an Illumina Solexa sequencer. After the
sequence reads had passed the quality check, the ELAND program was used
to align the reads to the hg18 provided by UCSC as part of the Illumina
data analysis pipeline. This was also done by the same company.
Thereafter the aligned sequence tags were fed into the FindPeaks
3.1.9.2. package written by the Canada Michael Smith Genome Sciences
Centre in order to 1. analyze the short-read sequences to identify areas
of enrichment of the protein of interest and 2. generating wig files for
use in the UCSC genome browser. The company performed a FindPeaks run
with standard settings. After I adjusted the settings to my own wished,
I performed another FindPeaks run. The wig file that was generated by
the company could be put into the genome browser without any problems.
After uploading the wig file created by myself, it seems at first that
Manage Custom Tracks
genome: Human assembly: Mar. 2006 [hg18]
Name
Description
Type
Doc
test3_duplicates_subpeaks_len_triangle
<http://genome.ucsc.edu/cgi-bin/hgCustom?hgsid=114649973&hgct_table=ct_t
est3duplicatessubpeakslentriangle>
test3_ht:1.0_FL:triangle_dupe_rds_inc
wiggle_0
But when I want to go to the genome browser, I the following error
removing custom tracks
Do you know by any chance what went wrong? I've already tried the
standard FindPeaks settings the company also used, but it doesn't work
form me.
I hope someone will have an answer for me. Thanks in advance!
Regards,
Marije
Drs. Marije Telgenkamp, MD
PhD candidate Molecular Cell Biology
Department of Science and Technology
University of Twente
Building Zuidhorst, room ZH125
PO box 217
7500 AE Enschede, The Netherlands
Phone: [phone]
Fax: [phone]

_______________________________________________
Genome maillist - ***@soe.ucsc.edu
http://www.soe.ucsc.edu/mailman/listinfo/genome