Soap can't scaffold !

[macb]

HI everyone. I'm new here. I got a problem blocked me for days and plz help me.
I'm using Soap to assemble de novo a genome with an estimated size of 2 GB.

Command :
SOAPdenovo127mer all -p 60 -s $conffile -o $dir/soapdenovo.assembly -K $nbround -R

However soapdenovo.assembly.scaf and soapdenovo.assembly.scaf_gap are all size 0.

I tested several Kmers from 30 to 70 and I have 4 libraries:
1 Pair End (100pb) + 2 Mate Pairs 100pb + 2 Mate Paris 50 pb
All worked correctly till scaffolding which is doing nothing.

Below my config_file details :
max_rd_len=100
[LIB]
avg_ins=453
reverse_seq=0
asm_flags=3
rank=0
pair_num_cutoff=3
q1=/home1/camine/work/SOAP/Pair_END/P34female.f.fq
q2=/home1/camine/work/SOAP/Pair_END/P34female.r.fq

[LIB]
avg_ins=5000
reverse_seq=1
asm_flags=2
rank=0
q1=/home1/camine/work/SOAP/Mate_Pair/GZ1/R1.NotEmpty.NotLink.fastq
q2=/home1/camine/work/SOAP/Mate_Pair/GZ1/R2.NotEmpty.NotLink.fastq

[LIB]
avg_ins=5000
reverse_seq=1
asm_flags=2
rank=0
q1=/home1/camine/work/SOAP/Mate_Pair/GZ2/R1.NotEmpty.NotLink.fastq
q2=/home1/camine/work/SOAP/Mate_Pair/GZ2/R2.NotEmpty.NotLink.fastq

[LIB]
avg_ins=5000
reverse_seq=1
asm_flags=2
rank=1
q1=/home1/camine/work/SOAP/Mate_Pair/GZ1/R1.NotEmpty.LinkerTrimmed.fastq
q2=/home1/camine/work/SOAP/Mate_Pair/GZ1/R2.NotEmpty.LinkerTrimmed.fastq

[LIB]
avg_ins=5000
reverse_seq=1
asm_flags=2
rank=1
q1=/home1/camine/work/SOAP/Mate_Pair/GZ2/R1.NotEmpty.LinkerTrimmed.fastq
q2=/home1/camine/work/SOAP/Mate_Pair/GZ2/R2.NotEmpty.LinkerTrimmed.fastq


Any one who have succeed? Thank you in advance.

[jebowers]

Version r235 was having problems seeing sequences from even numbered ranks in scaffolding. I do not know how this problem was impacted by rank 0. This problem was apparently fixed in the current version.

I would give it a try with ranks 1 and 2 instead of 0 and 1, and install the current version of SOAPdenovo2.

[macb]

Hi jebowers !
Thank tou for your response

I trimmed firstly my read at the end both Pair end and Mate Pair. Then I used the current version of SOAPdenovo2 with ranks 1 and 2.

It worked correctly just before sacffoolding. These files are still size 0 :

soapdenovo.assembly.links
soapdenovo.assembly.scaf_gap
soapdenovo.assembly.scaf
soapdenovo.assembly.gapSeq
soapdenovo.assembly.contigPosInscaff.


In the config file I have well precised reverse_seq=0 for Pair End and reverse_seq=1 for Mate Pair. However the LogSoap file show as that links were incorrectly oriented :


For insert size: 443
Total PE links 334538652
Normal PE links on same contig 0
Incorrect oriented PE links 334538652
PE links of too small insert size 0
PE links of too large insert size 0
Correct PE links 0
Accumulated connections 0
Use contigs longer than 443 to estimate insert size:
PE links 0
Too few PE links.
0 new connections.

For insert size: 5000
Total PE links 4798230
Normal PE links on same contig 0
Incorrect oriented PE links 4798230
PE links of too small insert size 0
PE links of too large insert size 0
Correct PE links 0
Accumulated connections 0
Use contigs longer than 5000 to estimate insert size:
PE links 0
Too few PE links.
0 new connections.

[macb]

Hi jebowers !
Thank tou for your response

I trimmed firstly my read at the end both Pair end and Mate Pair. Then I used the current version of SOAPdenovo2 with ranks 1 and 2.

It worked correctly just before sacffoolding. These files are still size 0 :

soapdenovo.assembly.links
soapdenovo.assembly.scaf_gap
soapdenovo.assembly.scaf
soapdenovo.assembly.gapSeq
soapdenovo.assembly.contigPosInscaff.


In the config file I have well precised reverse_seq=0 for Pair End and reverse_seq=1 for Mate Pair. However the LogSoap file show as that links were incorrectly oriented :


For insert size: 443
Total PE links 334538652
Normal PE links on same contig 0
Incorrect oriented PE links 334538652
PE links of too small insert size 0
PE links of too large insert size 0
Correct PE links 0
Accumulated connections 0
Use contigs longer than 443 to estimate insert size:
PE links 0
Too few PE links.
0 new connections.

For insert size: 5000
Total PE links 4798230
Normal PE links on same contig 0
Incorrect oriented PE links 4798230
PE links of too small insert size 0
PE links of too large insert size 0
Correct PE links 0
Accumulated connections 0
Use contigs longer than 5000 to estimate insert size:
PE links 0
Too few PE links.
0 new connections.

In fact my reads were not well oriented. I have done a mistake in my script for keeping pair end reads. Now after fixing that, Soap is scaffolding correctly.