[danix]

Hi boetsie, thanx again for your quick reply.
Here is a part of my .contig file. It was created by ace2contig (AMOS pack) and the input was the .ace that phrap generated after the assembly.
I'll try to use the script u attached.
Thank you so much again!

##Contig1 1 458 bases, 00000000 checksum.
agttcggcatggggtcaggtggttccactgcgctattgccgccaggcaaattcttcaatc
tgagaaagctgatgtaagtaattcgttcattcgctacaaggccagaaacacttcttgggt
gttgtatggttaagcctcacgggtaattagtatgggttagctcaacgtatcgctacgctt
acacaccccacctatcaacgttgtggtctccaacggccctttaggaccctcaaggggtca
gggatgactcatctcagggctcgcttcccgcttagatgctttcagcggttatcgattccg
aacttagctaccgggcagtgccactggcgtgacaacccgaacaccagaggttcgttcact
ccggtcctctcgtactaggagcaactcccttcaatcatccaacgcccacggcagataggg
accgaactgtctcacgacgttctgaacccagctcgcgt
#FZ92HC101BPK62(0) [] 458 bases, 00000000 checksum. {1 458} <1 459>
agttcggcatggggtcaggtggttccactgcgctattgccgccaggcaaattcttcaatc
tgagaaagctgatgtaagtaattcgttcattcgctacaaggccagaaacacttcttgggt
gttgtatggttaagcctcacgggtaattagtatgggttagctcaacgtatcgctacgctt
acacaccccacctatcaacgttgtggtctccaacggccctttaggaccctcaaggggtca
gggatgactcatctcagggctcgcttcccgcttagatgctttcagcggttatcgattccg
aacttagctaccgggcagtgccactggcgtgacaacccgaacaccagaggttcgttcact
ccggtcctctcgtactaggagcaactcccttcaatcatccaacgcccacggcagataggg
accgaactgtctcacgacgttctgaacccagctcgcgt
##Contig2 1 379 bases, 00000000 checksum.
ttctgagggaacacgcgttctgcgcgggttgtcttggtgctcactgttttccgccccgga
gtttgtggggtgttgggggtggtgggtgtgtgttgtttgagaagtgcatagtggatgcga
gcatctagcccggcgagttccttggtgttcttgttgggttgtgtgttctgcaatttcgat
tctggtttgtgcgatcgcgtgttgtgatcgttgatttttgtttgttgtccgcattcgcgt
ctcgggcactgtttggtgtgtggggtgtgtttgtgggtgttgttgtaagtgtttgagggc
gttcggtggatgccttggtaccaggagccgatgaaggacggccgtgcggtgggtcagtga
taaatcgacatgttaggtg
#FZ92HC101BFQDN(0) [] 379 bases, 00000000 checksum. {1 379} <1 380>
ttctgagggaacacgcgttctgcgcgggttgtcttggtgctcactgttttccgccccgga
gtttgtggggtgttgggggtggtgggtgtgtgttgtttgagaagtgcatagtggatgcga
gcatctagcccggcgagttccttggtgttcttgttgggttgtgtgttctgcaatttcgat
tctggtttgtgcgatcgcgtgttgtgatcgttgatttttgtttgttgtccgcattcgcgt
ctcgggcactgtttggtgtgtggggtgtgtttgtgggtgttgttgtaagtgtttgagggc
gttcggtggatgccttggtaccaggagccgatgaaggacggccgtgcggtgggtcagtga
taaatcgacatgttaggtg

[danix]

Hi, I forgot to mention that I also have the .sff if I can use them to create .mates it'll be great.
Can I? If so, how?

[boetsie]

Hi, I forgot to mention that I also have the .sff if I can use them to create .mates it'll be great.
Can I? If so, how?

I have no idea... I've never used a .sff file. How does it look like? why do you want to use it, does it contain additional data?

If the mates that are present in the .contig file, are all present in the two .fasta files, you can just use the two fasta files to create the .mates file.

[danix]

Hi, the 454 output is sff (looks like a binary file), but we use a script called sff_extract to convert this data in fasta, xml and quality files. I was just reading now that "The 454 paired-end protocol will generate reads which contain the forward and reverse direction in one read, separated by a linker."
So I think the key to generate .mates is .sff, but I don't know how.
I think I shouldn't be so complicated...

[danix]

I have no idea... I've never used a .sff file. How does it look like? why do you want to use it, does it contain additional data?

If the mates that are present in the .contig file, are all present in the two .fasta files, you can just use the two fasta files to create the .mates file.

How do I create the .mates? I tried with the script u send me and the output isn't fine. Besides I don't understand why FZ92HC101CZUHH.1 and FZ92HC102IDBLW.2 are in the same line. How can I tell that they are mates? I'm really lost and confused now...

FZ92HC101CZUHH.1 FZ92HC102IDBLW.2 libname
FZ92HC101DJEHD.1 FZ92HC102JYG94.2 libname
FZ92HC101DUWKQ.1 FZ92HC102HS1LU.2 libname
FZ92HC101CUUV5.1 FZ92HC102G8H4Z.2 libname
FZ92HC101EMKQX.1 FZ92HC102HOD38.2 libname
FZ92HC101CE653.1 FZ92HC102HO0J7.2 libname
FZ92HC101ECTBB.1 FZ92HC102IBNJJ.2 libname
FZ92HC101DXMSC.1 TGATCCGGCGCAGGCGTATCTGGGCTCGGATCGTGCCTGGTGCCGACGGCGATGAACGAC
libname
FZ92HC101C587C.1 FZ92HC102F3E16.2 libname
FZ92HC101BZ63S.1 CGGTCGGCCGCGGCCGATCTCGGGATTGCGCGGCGTGTGCAT
libname
FZ92HC101DEODE.1 CCGCGTGGACATGCCGTTCGAGGAACCGTGGACGCAACC
libname
FZ92HC101DP9HX.1 ATCGGCTATGCACAGGTCATCGAGTATCTCGACGGCG
libname
FZ92HC101EE90B.1 ACGTCCGACGTGATCAGGAGCGAGTCGGTGACGGCGCTTCGCACTCCGAGGG
libname
TTTGATGATCGACATCAAT GCGTTCGACTACCAGTTCGTCGGACCATCCGGGTAGCGTGTCGCAAGGGTCGGTTCCGAA
libname
CGTTCGCTGAGCACCGCCGAATCGAGCAGTTCGCGGATCTCGTCGAACGTCCNCGA FZ92HC102GE3MB.2 libname
CGTACGGATGTAGCTGGTGAAGAGGTCCCTTGCGGGCGGAGAAGTCGAGTCGTTCCGTCG TCGAGAGGCCGCGGAAGCGGCCGGAAAGGACGGCAACGATGTTTGACCGTTTCAACTCAG
libname
FZ92HC101DBOTK.1 FZ92HC102GVOHT.2 libname
FZ92HC101BEEQB.1 TCTGCGTGGAGACCGTGACGGCTGATCTACGGCCNCCTCGGCCGATGATCGCCGCCT

[catfisher]

Bambus error: library priority

Boetsie and danix, I noticed that you may do a lot of work using Bambus, I also get the contigs generated from CLCbio. I know how to get the .contig file for Bambus, and I also got a mates file following your instructions, but when I rum goBambus, I got an error:
20100710|193857| 16658| Grommit(/home/aubsxl/bin/bambus/bin/grommit -i ctg2660_BES_mapping_704.inp -o ctg2660_BES_
mapping_704.out.xml -C ctg2660_BES_mapping_704.grommit.conf --append --logfile goBambus.log --debug 1) script fail
ed
20100710|204158|24277|grommit|FATAL|9: Priority not specified: at least one library must be assigned a priority

I don't know what's the 'priority', how can I do to solve this problem? could you all give any help? Thanks in advance.

[boetsie]

Hi catfisher,

i´ve had this error too. To solve it, you should set a priority in the .conf file. A file named default.conf is generated once you have run Bambus. This file contains the default parameters. Change or edit the line to;

priority ALL 1

to the file.
If you did not run Bambus yet, you should create one from scratch. See the below links for more information. Once you have the .conf file, you should add it to the command line options with for example;
goBambus -c test.contig -m test.mates -C default.conf -o test-bambus

For more information about the .config file see;

http://sourceforge.net/apps/mediawiki/amos/index.php?title=Bambus_Manual#The_configuration_file

For an example see;

http://sourceforge.net/apps/mediawiki/amos/index.php?title=Bambus.conf

Marten

[catfisher]

grommit script failed

Marten, thanks for your quick reply. I editted my configure file as you suggested and run goBambus again, but still failed.
I used the .conf as:
# Priorities
priority ALL 1
# The following lines can be un-commented to specify certain
# per-library settings

# Redundancies
# redundancy lib_some 1

# allowed error
# error MUMmer 0.5

# overlaps allowed
# overlaps MUMmer Y

# Global redundancy
redundancy 2

# min group size
mingroupsize 0

The log information for goBambus is :
Parsing links out of input file
Step 100: running detective
Combining XML files
Step 200: making the xmls
starting
Done
Step 300: Preparing contig links
starting
Done
Step 400: Running scaffolder
Grommit(/home/aubsxl/bin/bambus/bin/grommit -i ctg2660_BES_mapping_704.inp -o ctg2660_BES_mapping_704.out.xml -C c
tg2660_BES_mapping_704.grommit.conf --append --logfile goBambus.log --debug 1) script failed

The error information from goBambus.error file is:
20100712|123807| 10451| Grommit(/home/aubsxl/bin/bambus/bin/grommit -i ctg2660_BES_mapping_704.inp -o ctg2660_BES_
mapping_704.out.xml -C ctg2660_BES_mapping_704.grommit.conf --append --logfile goBambus.log --debug 1) script fail
ed

The first several lines from my mates files is:
library libname 200 500
HWUSI-EAS1665_0002:2:1:1022:18088#0/1 HWUSI-EAS1665_0002:2:1:1022:18088#0/2 libname
HWUSI-EAS1665_0002:2:1:1029:11872#0/1 HWUSI-EAS1665_0002:2:1:1029:11872#0/2 libname
HWUSI-EAS1665_0002:2:1:1029:11034#0/1 HWUSI-EAS1665_0002:2:1:1029:11034#0/2 libname
HWUSI-EAS1665_0002:2:1:1030:19457#0/1 HWUSI-EAS1665_0002:2:1:1030:19457#0/2 libname
HWUSI-EAS1665_0002:2:1:1031:12133#0/1 HWUSI-EAS1665_0002:2:1:1031:12133#0/2 libname

Marten, could you look at these information and point out what's wrong with this? I have no idea. Thanks a lot,

Kevin

[boetsie]

Hi catfisher,

hmmm weird error, since it doesn't point out where it goes wrong. Is that the only error?

Some thing that might help;

replace ":" and "#" in the readnames to underscores ("_"). E.g.;

HWUSI-EAS1665_0002:2:1:1022:18088#0/1
will be;
HWUSI-EAS1665_0002_2_1_1022_18088_0/1

do this both in the .mates file and .contig file.

Code to do this is;

cat input.mates | sed s/#/_/g | sed s/:/_/g > output.mates

where input.mates is the input file, and output.mates the converted output file.

I don't know if this really works...

Otherwise it might be a good idea to contact Bambus developers, since i'm not to familiar with Bambus.

Good luck.

Cheers,
Marten

[themerlin]

Catfisher,

I had the same error months ago. I ended up filtering my contigs so I only kept longer contigs (>500nts) with high coverage (depends on your dataset). I didn't change my mates file and then it suddenly worked. I'm not quite sure why, but it might be worth a shot.

Jason

[catfisher]

Catfisher,

I had the same error months ago. I ended up filtering my contigs so I only kept longer contigs (>500nts) with high coverage (depends on your dataset). I didn't change my mates file and then it suddenly worked. I'm not quite sure why, but it might be worth a shot.

Jason

I headed 100k lines of the contig and mates files and rerun the program for these data, the program also worked now.
Does anyone know how much data size we can handle with the bambus? I am afraid that it has a built-in limit for how big the input data can be input. I have 704 contigs in the .contig file and 3434936 x2 paired ends, the program didn't work if I loaded all of them. I tested one with contigs less than 500bp (some are about 200bp), it worked also. How big were the input data when you all used the Bambus? Thanks,

Kevin

[boetsie]

This thread was solved by a program developed by myself which can scaffold assembled contigs in .fasta format with paired-end and/or mate pair sequences. No conversion of file formats are required. See this thread;

http://seqanswers.com/forums/showthread.php?t=4124

[seb567]

You can try SSPACE too. It is a scaffolder for next-gen data.

Bioinformatics paper:

http://bioinformatics.oxfordjournals.org/content/early/2010/12/12/bioinformatics.btq683.short

SEQanswers thread:

http://seqanswers.com/forums/showthread.php?t=8350

Download:

http://www.baseclear.com/sequencing/data-analysis/bioinformatics-tools/sspace/

-seb

[boetsie]

You can try SSPACE too. It is a scaffolder for next-gen data.

-seb

Hmmm, that is the program I meant, since I'm the developer haha. I refered to the wrong link in my previous reply

Boetsie

[seb567]

Hmmm, that is the program I meant, since I'm the developer haha. I refered to the wrong link in my previous reply

Boetsie

That's funny !