Also attached is a sample output (which I've truncated below)
1 >Cluster 0 & bsp;
2 0 271623nt, >19_contig-1000000... at +/100.00%
3 1 63731nt, >19_contig-3... at +/100.00%
4 2 346149nt, >21_contig-8000003... *
5 3 243006nt, >23_contig-1000002... at +/100.00%
6 4 330249nt, >25_contig-9000002... at +/100.00%
7 5 63731nt, >27_contig-1000002... at +/100.00%
8 6 271674nt, >27_contig-7000002... at -/99.99%
9 7 63749nt, >29_contig-4000000... at +/100.00%
10 8 242978nt, >29_contig-11000002... at +/100.00%
We'd hope not to have multiple entries for the kmer in a cluster.
Hope this is useful,
Regards
Mitch
On Fri, May 25, 2012 at 11:35 AM, Mitchell Stanton-Cook
<[email protected] <mailto:[email protected]>> wrote:
Hi Seb,
Thank you. We're still doing some more work this end. We
really really really like Ray! We are more than happy to work
through these problems.
We hope to be pushing <information redacted> through Ray in a
few months. We just want to be sure that we are getting
useful results.
If there is anything else we can do to help/bugfix please do
not hesitate to let us know.
Regards
Mitch
On Fri, May 25, 2012 at 11:23 AM, Sébastien Boisvert
<[email protected]
<mailto:[email protected]>> wrote:
Just to let you know that I may have found the problem.
See https://github.com/sebhtml/ray/issues/55
Sébastien
________________________________________
De : Mitchell Stanton-Cook [[email protected]
<mailto:[email protected]>]
Date d'envoi : 17 mai 2012 22:32
À : Sébastien Boisvert
Objet : Re: Duplicate sequence in contigs
Hi Seb,
Thanks for the reply.
A bit more background:
-------------------------------
This is 100 bp Illumina PE data (insert of ~300 bp s.d of
about ~10%).
~ 1000X coverage. Ray (1.7) did not like this high
coverage (you have previously commented on the user-list
about this). We sampled this down to ~100X coverage.
We also cleaned the reads (you have pointed this is not
necessary, but having a consistent cleaned set makes
downstream analysis i.e. snp calling much easier). All
input bases have a Q score >= 30. Reads > 70bp after
trimming were filtered. After this the mean reads size is
~97 bp. We only consider read pairs (if 1 one of the
sequences in the pairs fails a cleaning criterion, both
do) and hence no single end reads go into Ray.
Ray was executed like this (we used Ray's internal
estimations/calculations to determine the best parameters):
mpiexec -n $PROC Ray -i XXXX.fastq -k $K -o $OUT
We looked at kmers from 15-35 in increments of 2.
The genome is ~1.8 Mb. From the assemblies it appears
there are not a lot repetitive elements in the genome. I
have attached a csv of the results (XXXX_ALL.csv) (
abbreviations: c= contigs, s= scaffold, N = scaffolding
character).
Results for kmer 21 and kmer 23 are interesting.
kmer 21) As previously mentioned we have a 63 Kb
duplication. For 63 Kb the start of the two contigs are
almost identical (3 mismatches and 1 gap):
Score = 1.165e+05 bits (63086), Expect = 0.0
Identities = 63093/63096
<tel:63093%2F63096><tel:63093%2F63096> (99%), Gaps =
1/63096 (0%)
Strand=Plus/Plus
I have attached the alignment (XXXX_b2s.aln)
There is also a smaller 5 Kb duplication detected:
Score = 9583 bits (5189), Expect = 0.0
Identities = 5192/5193 (99%), Gaps = 1/5193 (0%)
Strand=Plus/Minus
kmer 23) (Notice there is about 63 Kb difference between
the "c 100 bp total len" in the .csv file of kmer 21 and
kmer 23). From the blast2seq there is no such 63 Kb
duplication found.
Now, once again focusing on the "c 100 bp total len" in
the .csv file. I propose kmer 21 duplicate, kmer 23
no-duplicate, kmer 25 non-duplicate, kmer 27 duplicate (I
verified this true), kmer 29 non-duplicate. Strangely
kmer 31 is missing about 200 Kb in comparison to the
other assemblies.
What we are wondering is:
1) Why is there large, almost identical duplicates
present in the assemblies?
2) Why do we see it in some kmers and not others? (I
could understand if lower kmer = duplicates, higher kmers
= non-duplicates if these duplicates are propagated by a
sequencing error)
3) Is this a bug or an inherit issue with graph based
assembly?
4) If possible, how can we fix this?
5) How can I help you with this?
I hope I have provided you with enough information. If
not please let me know.
p.s. I have know problem with this going to the list as
long as the attachments are not included.
Regards
Mitch
On Fri, May 18, 2012 at 12:25 AM, Sébastien Boisvert
<[email protected]
<mailto:[email protected]><mailto:[email protected]
<mailto:[email protected]>>> wrote:
Hi,
Is the 63 kB perfectly duplicated in your assembly ?
Le 2012-05-17 01:26, Mitchell Stanton-Cook a écrit :
Hi Seb,
Hope all is well.
I was wondering if you have ever seen duplicate sequence
in contigs.
We have ~63 kB duplicate at the start of two different
contigs. Beyond this it's unique.
This is Ray 1.7.
I'm re-running with Ray2.0rc5.
I came across these posts (ABySS specific):
http://groups.google.com/group/abyss-users/browse_thread/thread/264e894b4ec0c96d/30dab75afa686878
http://groups.google.com/group/abyss-users/browse_thread/thread/7a03ee033b11afc4
http://groups.google.com/group/abyss-users/browse_thread/thread/f0f3a650bd12cf1e
Any ideas?
Regards
Mitch
