Question: MapSplice Error: filter alignment failed
0
gravatar for liguanghaoli
5 months ago by
liguanghaoli0 wrote:

Hi, When I use MapSplice to do mapping in RNA seq data analysis. The error occurred and there was no bam file generated. The error information is listed as follow:

[Thu Jan 11 15:42:04 2018] Inspecting Bowtie index files
[Thu Jan 11 15:42:04 2018] Checking reference sequence length
[Thu Jan 11 15:42:29 2018] Checking consistency of Bowtie index and reference sequence
[Thu Jan 11 15:42:29 2018] Checking read format
[Thu Jan 11 15:44:54 2018] Running MapSplice multi-thread
[Thu Jan 11 17:42:09 2018] Generating junctions from sam file
[Thu Jan 11 17:43:07 2018] Filtering junction by min mis and min lpq
[Thu Jan 11 17:43:07 2018] Filtering junction by ROC argu noncanonical
[Thu Jan 11 17:43:08 2018] Generating synthetic junction sequences
[Thu Jan 11 17:43:27 2018] Checking Bowtie index files
[Thu Jan 11 17:43:27 2018] Building Bowtie index for junction synthetic sequence
[Thu Jan 11 17:43:47 2018] Running MapSplice multi-thread
[Thu Jan 11 20:25:13 2018] Running alignment handler
[MapSplice Running Failed]
Error: filter alignment failed

Is there anyone have encounted this error before? I have read the user guide of MapSplice and couldn't find the solution. I think there may be some problems in filtering unmapped reads but don't know how to solve it. Could you please help me? Thank you very much. Guanghao Li

rna-seq • 337 views
ADD COMMENTlink modified 4 months ago by simo.aufiero0 • written 5 months ago by liguanghaoli0

maybe you should send the data to mapsplice at netlab.uky.edu, they can help you

ADD REPLYlink written 4 months ago by staypositive0
0
gravatar for simo.aufiero
5 months ago by
European Union
simo.aufiero0 wrote:

Hi Guanghao Li, I had the same problem. I am running mapsplice using the genome reference mm10. I did the analysis in the past by using mm9 and mapsplice pipeline finished without any problem with that genome reference. What genome reference are you using?

Simona

ADD COMMENTlink written 5 months ago by simo.aufiero0

Hi, Simona, I use hg19 as reference. And now I run MapSplice in another computer, the program work well. So I'm also comfused about the error.

ADD REPLYlink modified 5 months ago • written 5 months ago by liguanghaoli0
0
gravatar for spin
4 months ago by
spin0
spin0 wrote:

Hello, same error message here but it is inconsistent. Some of my samples from the same batch run perfectly fine. I use GRCh38 as reference. Have you been able to figure out the problem?

Thanks J

ADD COMMENTlink written 4 months ago by spin0
0
gravatar for simo.aufiero
4 months ago by
European Union
simo.aufiero0 wrote:

Hi Spin, this is the command line that I used to perfom the analysis: python mapsplice.py -p 16 --bam -o mapsplice_out001 -c Chromosomes -x BowtieIndex/genome -1 paired-001.R1.fastq -2 paired-001.R2.fastq --gene-gtf gencode.vM16.annotation.gtf --fusion-non-canonical --filtering 1 --min-map-len 25 --min-fusion-distance 200

Below I reported the log file of the analysis. As you can see in my case the error generated when mapsplice tried to build bowtie index for fusion junction synthetic sequence. I sent an email to netlab (mapsplice@netlab.uky.edu) asking how I could solve the problem. They suggested to reduce the false positives: 1. by increasing the --min-map-len 2. if 1 does not solve the problem, by turning off --fusion-non-canonical

By increasing the --min-map-len to 50 and by turning off --fusion-non-canonical the analysis finished without any problem. I hope this can help you.

Simona

[Tue Jan 30 08:51:54 2018] Beginning Mapsplice run (MapSplice v2.2.0) [Tue Jan 30 08:51:54 2018] Bin directory: /scratch/1055713.batch1.lisa.surfsara.nl/MS22_mm10/bin/ [Tue Jan 30 08:51:54 2018] Preparing output location mapsplice_out002/ [Tue Jan 30 08:51:54 2018] Checking files or directory: paired-002.R1.fastq [Tue Jan 30 08:51:54 2018] Checking files or directory: paired-002.R2.fastq [Tue Jan 30 08:51:54 2018] Checking files or directory: Chromosomes/ [Tue Jan 30 08:51:54 2018] Checking Bowtie index files [Tue Jan 30 08:51:54 2018] Inspecting Bowtie index files [Tue Jan 30 08:51:54 2018] Checking reference sequence length [Tue Jan 30 08:52:15 2018] Checking consistency of Bowtie index and reference sequence [Tue Jan 30 08:52:15 2018] Checking read format -----[Read Format: FASTQ] -----[Read Type: Pair End] -----[Total # Reads: 134960420] -----[Max Read Length: 151] -----[Min Read Length: 50] -----[Max Quality Score: 74] -----[Min Quality Score: 35] -----[Quality Score Scale: Phred+33] [Tue Jan 30 09:08:00 2018] Running MapSplice multi-thread [Tue Jan 30 10:44:48 2018] Generating junctions from sam file [Tue Jan 30 10:46:28 2018] Filtering junction by min mis and min lpq [Tue Jan 30 10:46:31 2018] Filtering junction by ROC argu noncanonical [Tue Jan 30 10:46:33 2018] Generating synthetic junction sequences [Tue Jan 30 10:46:52 2018] Checking Bowtie index files [Tue Jan 30 10:46:52 2018] Building Bowtie index for junction synthetic sequence [Tue Jan 30 10:47:21 2018] Running MapSplice multi-thread [Tue Jan 30 14:33:45 2018] Running alignment handler [Tue Jan 30 15:03:37 2018] Parsing cluster regions [Tue Jan 30 15:03:51 2018] Clustering regions [Tue Jan 30 15:03:57 2018] Running MapSplice multi-thread fusion [Tue Jan 30 16:32:55 2018] Generating fusion junctions from sam file and filter by anchor [Tue Jan 30 17:54:46 2018] Filtering original fusion junction [Tue Jan 30 17:57:56 2018] Synthetic fusion junctions sequence [Wed Jan 31 14:26:09 2018] Checking Bowtie index files [Wed Jan 31 14:26:09 2018] Building Bowtie index for fusion junction synthetic sequence Error: Reference sequence has more than 2^32-1 characters! Please divide the reference into batches or chunks of about 3.6 billion characters or less each and index each independently. Command: /scratch/1055713.batch1.lisa.surfsara.nl/MS22_mm10/bin/bowtie-build mapsplice_out002/tmp/fusion/fusion_synthetic_sequence.txt mapsplice_out002/tmp/fusion/syn_fusion_idx_prefix [MapSplice Running Failed] Error: Splice sequence indexing failed cp: cannot stat 'circular_RNAs.txt': No such file or directory cp: cannot stat 'stats.txt': No such file or directory

ADD COMMENTlink written 4 months ago by simo.aufiero0
Please log in to add an answer.

Help
Access

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.
Powered by Biostar version 2.3.0
Traffic: 1003 users visited in the last hour