mstubb
commented
6 years ago Hi Linda,
Have you increased the memory available to your docker image? Have a look at the last paragraph in the docker section of the readme: https://github.com/teichlab/bracer#docker-image https://github.com/teichlab/bracer#docker-image
Let us know if that helps.
Mike
On 9 Aug 2018, at 01:34, Linda-Lan notifications@github.com wrote:
Hi bracer team,
I set up bracer through downloading the docker image directly, but ran error with test data as following. Do you have any suggestions for it?
Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer assemble -p 4 cell1 /scratch/out_test2 /scratch/cell1_1.fastq /scratch/cell1_2.fastq
Trimming raw reads
Detecting installed version of Cutadapt: 1.14 Trimming completed
Finding recombinant-derived reads
Attempting new assembly for ['BCR_H', 'BCR_K', 'BCR_L']
Detected average R1 read length: 50.0 Short read length detected. BraCeR will run two rounds of alignment for heavy chain
BCR_H
56433 reads; of these: 56433 (100.00%) were paired; of these: 34987 (62.00%) aligned concordantly 0 times 21446 (38.00%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
34987 pairs aligned concordantly 0 times; of these: 76 (0.22%) aligned discordantly 1 time
34911 pairs aligned 0 times concordantly or discordantly; of these: 69822 mates make up the pairs; of these: 69305 (99.26%) aligned 0 times 248 (0.36%) aligned exactly 1 time 269 (0.39%) aligned >1 times 38.60% overall alignment rate 56433 reads; of these: 56433 (100.00%) were paired; of these: 55425 (98.21%) aligned concordantly 0 times 1008 (1.79%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
55425 pairs aligned concordantly 0 times; of these: 20563 (37.10%) aligned discordantly 1 time
34862 pairs aligned 0 times concordantly or discordantly; of these: 69724 mates make up the pairs; of these: 69316 (99.41%) aligned 0 times 379 (0.54%) aligned exactly 1 time 29 (0.04%) aligned >1 times 38.59% overall alignment rate
BCR_K
56433 reads; of these: 56433 (100.00%) were paired; of these: 48166 (85.35%) aligned concordantly 0 times 8267 (14.65%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
48166 pairs aligned concordantly 0 times; of these: 112 (0.23%) aligned discordantly 1 time
48054 pairs aligned 0 times concordantly or discordantly; of these: 96108 mates make up the pairs; of these: 95668 (99.54%) aligned 0 times 187 (0.19%) aligned exactly 1 time 253 (0.26%) aligned >1 times 15.24% overall alignment rate
BCR_L
56433 reads; of these: 56433 (100.00%) were paired; of these: 31447 (55.72%) aligned concordantly 0 times 24986 (44.28%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
31447 pairs aligned concordantly 0 times; of these: 268 (0.85%) aligned discordantly 1 time
31179 pairs aligned 0 times concordantly or discordantly; of these: 62358 mates make up the pairs; of these: 61680 (98.91%) aligned 0 times 134 (0.21%) aligned exactly 1 time 544 (0.87%) aligned >1 times 45.35% overall alignment rate
Assembling Trinity Contigs
BCR_H
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases https://github.com/trinityrnaseq/trinityrnaseq/releases Wednesday, August 8, 2018: 23:45:43 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:45:44 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:45:44 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H Wednesday, August 8, 2018: 23:45:44 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:45:44 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:45:44 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:45:44 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa Wednesday, August 8, 2018: 23:45:44 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf --------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.tmp Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:45:46 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.finished -------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/filter_iworm_by_min_length_or_cov.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa 100 10 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/inchworm.K25.L25.DS.fa.min100 Running CMD: bowtie2-build -o 3 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/inchworm.K25.L25.DS.fa.min100 1>/dev/null Running CMD: bash -c " set -o pipefail;bowtie2 --local -k 2 --threads 4 -f --score-min G,46,0 -x /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa | samtools view -@ 4 -F4 -Sb - | samtools sort -m 134217728 -@ 4 -no - - > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/iworm.bowtie.nameSorted.bam" Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/scaffold_iworm_contigs.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/iworm.bowtie.nameSorted.bam /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/iworm_scaffolds.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/GraphFromFasta -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa -r /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa -min_contig_length 200 -min_glue 2 -glue_factor 0.05 -min_iso_ratio 0.05 -t 4 -k 24 -kk 48 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/iworm_cluster_welds_graph.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/BubbleUpClustering -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa -weld_graph /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/iworm_cluster_welds_graph.txt -min_contig_length 200 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/GraphFromIwormFasta.out Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/CreateIwormFastaBundle -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/GraphFromIwormFasta.out -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/bundled_iworm_contigs.fasta -min 200 Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/ReadsToTranscripts -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa -f /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/bundled_iworm_contigs.fasta -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/readsToComponents.out -t 4 -max_mem_reads 50000000 Running CMD: /usr/bin/sort --parallel=4 -T . -S 1G -k 1,1n /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/readsToComponents.out > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis/readsToComponents.out.sort Wednesday, August 8, 2018: 23:45:48 CMD: mkdir -p read_partitions/Fb_0/CBin_0 Wednesday, August 8, 2018: 23:45:49 CMD: touch partitioned_reads.files.list.ok Wednesday, August 8, 2018: 23:45:49 CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/write_partitioned_trinity_cmds.pl --reads_list_file partitioned_reads.files.list --CPU 1 --max_memory 1G --run_as_paired --seqType fa --trinity_complete --full_cleanup > recursive_trinity.cmds Wednesday, August 8, 2018: 23:45:49 CMD: touch recursive_trinity.cmds.ok Wednesday, August 8, 2018: 23:45:49 CMD: touch recursive_trinity.cmds.ok ------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:45:49 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 3 succeeded(3) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:45:57 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H.Trinity.fasta ###################################################################
BCR_K
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases https://github.com/trinityrnaseq/trinityrnaseq/releases Wednesday, August 8, 2018: 23:45:58 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:45:58 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:45:58 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K Wednesday, August 8, 2018: 23:45:58 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:45:58 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:45:58 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:45:58 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa Wednesday, August 8, 2018: 23:45:58 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf --------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.tmp Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:45:59 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.finished -------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/filter_iworm_by_min_length_or_cov.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa 100 10 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/inchworm.K25.L25.DS.fa.min100 Running CMD: bowtie2-build -o 3 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/inchworm.K25.L25.DS.fa.min100 1>/dev/null Running CMD: bash -c " set -o pipefail;bowtie2 --local -k 2 --threads 4 -f --score-min G,46,0 -x /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa | samtools view -@ 4 -F4 -Sb - | samtools sort -m 134217728 -@ 4 -no - - > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/iworm.bowtie.nameSorted.bam" Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/scaffold_iworm_contigs.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/iworm.bowtie.nameSorted.bam /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/iworm_scaffolds.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/GraphFromFasta -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa -r /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa -min_contig_length 200 -min_glue 2 -glue_factor 0.05 -min_iso_ratio 0.05 -t 4 -k 24 -kk 48 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/iworm_cluster_welds_graph.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/BubbleUpClustering -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa -weld_graph /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/iworm_cluster_welds_graph.txt -min_contig_length 200 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/GraphFromIwormFasta.out Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/CreateIwormFastaBundle -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/GraphFromIwormFasta.out -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/bundled_iworm_contigs.fasta -min 200 Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/ReadsToTranscripts -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa -f /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/bundled_iworm_contigs.fasta -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/readsToComponents.out -t 4 -max_mem_reads 50000000 Running CMD: /usr/bin/sort --parallel=4 -T . -S 1G -k 1,1n /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/readsToComponents.out > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis/readsToComponents.out.sort Wednesday, August 8, 2018: 23:46:01 CMD: mkdir -p read_partitions/Fb_0/CBin_0 Wednesday, August 8, 2018: 23:46:01 CMD: touch partitioned_reads.files.list.ok Wednesday, August 8, 2018: 23:46:01 CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/write_partitioned_trinity_cmds.pl --reads_list_file partitioned_reads.files.list --CPU 1 --max_memory 1G --run_as_paired --seqType fa --trinity_complete --full_cleanup > recursive_trinity.cmds Wednesday, August 8, 2018: 23:46:02 CMD: touch recursive_trinity.cmds.ok Wednesday, August 8, 2018: 23:46:02 CMD: touch recursive_trinity.cmds.ok ------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:46:02 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 1 succeeded(1) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:46:06 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K.Trinity.fasta ###################################################################
BCR_L
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases https://github.com/trinityrnaseq/trinityrnaseq/releases Wednesday, August 8, 2018: 23:46:07 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:46:07 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:46:07 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L Wednesday, August 8, 2018: 23:46:07 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:46:07 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:46:07 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:46:07 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa Wednesday, August 8, 2018: 23:46:07 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf --------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.tmp Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:46:09 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.finished -------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/filter_iworm_by_min_length_or_cov.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa 100 10 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/inchworm.K25.L25.DS.fa.min100 Running CMD: bowtie2-build -o 3 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/inchworm.K25.L25.DS.fa.min100 1>/dev/null Running CMD: bash -c " set -o pipefail;bowtie2 --local -k 2 --threads 4 -f --score-min G,46,0 -x /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/inchworm.K25.L25.DS.fa.min100 /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa | samtools view -@ 4 -F4 -Sb - | samtools sort -m 134217728 -@ 4 -no - - > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/iworm.bowtie.nameSorted.bam" Running CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/scaffold_iworm_contigs.pl /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/iworm.bowtie.nameSorted.bam /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/iworm_scaffolds.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/GraphFromFasta -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa -r /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa -min_contig_length 200 -min_glue 2 -glue_factor 0.05 -min_iso_ratio 0.05 -t 4 -k 24 -kk 48 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/iworm_cluster_welds_graph.txt Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/BubbleUpClustering -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa -weld_graph /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/iworm_cluster_welds_graph.txt -min_contig_length 200 > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/GraphFromIwormFasta.out Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/CreateIwormFastaBundle -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/GraphFromIwormFasta.out -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/bundled_iworm_contigs.fasta -min 200 Running CMD: /trinityrnaseq-Trinity-v2.4.0/Chrysalis/ReadsToTranscripts -i /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa -f /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/bundled_iworm_contigs.fasta -o /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/readsToComponents.out -t 4 -max_mem_reads 50000000 Running CMD: /usr/bin/sort --parallel=4 -T . -S 1G -k 1,1n /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/readsToComponents.out > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis/readsToComponents.out.sort Wednesday, August 8, 2018: 23:46:12 CMD: mkdir -p read_partitions/Fb_0/CBin_0 Wednesday, August 8, 2018: 23:46:12 CMD: touch partitioned_reads.files.list.ok Wednesday, August 8, 2018: 23:46:12 CMD: /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/write_partitioned_trinity_cmds.pl --reads_list_file partitioned_reads.files.list --CPU 1 --max_memory 1G --run_as_paired --seqType fa --trinity_complete --full_cleanup > recursive_trinity.cmds Wednesday, August 8, 2018: 23:46:13 CMD: touch recursive_trinity.cmds.ok Wednesday, August 8, 2018: 23:46:13 CMD: touch recursive_trinity.cmds.ok ------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:46:13 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 1 succeeded(1) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:46:21 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L.Trinity.fasta ###################################################################
Running BLAST
Performing Blast on ['BCR_H', 'BCR_K', 'BCR_L']
BCR_H
BCR_K
BCR_L
Running IgBLAST
Ig_seqtype: Ig Performing IgBlast on ['BCR_H', 'BCR_K', 'BCR_L']
BCR_H
BCR_K
BCR_L
START> MakeDBALIGNER> IgBlast ALIGNER_OUTPUT> cell1_BCR_H.fmt7 SEQ_FILE> cell1_BCR_H.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (3) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_H_db-pass.tab PASS> 1 FAIL> 2 END> MakeDb
START> MakeDBALIGNER> IgBlast ALIGNER_OUTPUT> cell1_BCR_K.fmt7 SEQ_FILE> cell1_BCR_K.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (1) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_K_db-pass.tab PASS> 1 FAIL> 0 END> MakeDb
START> MakeDBALIGNER> IgBlast ALIGNER_OUTPUT> cell1_BCR_L.fmt7 SEQ_FILE> cell1_BCR_L.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (1) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_L_db-pass.tab PASS> 1 FAIL> 0 END> MakeDb
Running Kallisto
Making Kallisto indices
[build] loading fasta file /scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.fa [build] k-mer length: 31 [build] warning: clipped off poly-A tail (longer than 10) from 1549 target sequences [build] warning: replaced 4 non-ACGUT characters in the input sequence with pseudorandom nucleotides [build] counting k-mers ... Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in load_entry_point('bracer==0.1', 'console_scripts', 'bracer')() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch Task().run() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 369, in run self.quantify(cell) File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 605, in quantify self.trimmed_fastq2, self.keep_trimmed_reads) File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/bracer_func.py", line 2110, in quantify_with_kallisto subprocess.check_call(index_command) File "/usr/lib/python3.5/subprocess.py", line 271, in check_call raise CalledProcessError(retcode, cmd) subprocess.CalledProcessError: Command '['/kallisto_linux-v0.43.1/kallisto', 'index', '-i', '/scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.idx', '/scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.fa']' returned non-zero exit status -9 Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd out_test Lindas-MacBook-Pro:out_test lindalan$ ls cellAAA Lindas-MacBook-Pro:out_test lindalan$ cd cellAAA/ Lindas-MacBook-Pro:cellAAA lindalan$ ls BLAST_output Trinity_output expression_quantification trimmed_reads IgBLAST_output aligned_reads filtered_BCR_seqs unfiltered_BCR_seqs Lindas-MacBook-Pro:cellAAA lindalan$ cd /Users/lindalan/docker/bracer/bracer/test_data Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 -g pdf /scratch/out_test2 usage: bracer [-h] [--ncores ] [--config_file
] [--resource_dir ] [--species SPECIES] [--loci LOCI [LOCI ...]] [--use_unfiltered] [--graph_format ] [--no_networks] [--IGH_networks] [--dist ] [--include_multiplets] [--infer_lineage] bracer: error: unrecognized arguments: -g /scratch/out_test2 Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 --g pdf /scratch/out_test2 Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in load_entry_point('bracer==0.1', 'console_scripts', 'bracer')() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch Task().run() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 746, in run self.loci, cells) File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 1036, in write_reconstruction_statistics pc = round((count/float(total_cells))100, 1) ZeroDivisionError: float division by zero Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 /scratch/out_test2 Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in load_entry_point('bracer==0.1', 'console_scripts', 'bracer')() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch Task().run() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 746, in run self.loci, cells) File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 1036, in write_reconstruction_statistics pc = round((count/float(total_cells))100, 1) ZeroDivisionError: float division by zero Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd out_test2 Lindas-MacBook-Pro:out_test2 lindalan$ ls cell1 cell2 cellA filtered_BCR_summary results Lindas-MacBook-Pro:out_test2 lindalan$ cd filtered_BCR_summary/ Lindas-MacBook-Pro:filtered_BCR_summary lindalan$ ls BCR_summary.txt Lindas-MacBook-Pro:filtered_BCR_summary lindalan$ cd .. Lindas-MacBook-Pro:out_test2 lindalan$ cd results/ Lindas-MacBook-Pro:results lindalan$ ls cell1 Lindas-MacBook-Pro:results lindalan$ cd cell1/ Lindas-MacBook-Pro:cell1 lindalan$ ls BLAST_output Trinity_output expression_quantification unfiltered_BCR_seqs IgBLAST_output aligned_reads filtered_BCR_seqs Lindas-MacBook-Pro:cell1 lindalan$ cd .. Lindas-MacBook-Pro:results lindalan$ ls cell1 Lindas-MacBook-Pro:results lindalan$ cd out -bash: cd: out: No such file or directory Lindas-MacBook-Pro:results lindalan$ cd .. Lindas-MacBook-Pro:out_test2 lindalan$ ls cell1 cell2 cellA filtered_BCR_summary results Lindas-MacBook-Pro:out_test2 lindalan$ cd .. Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd out_test Lindas-MacBook-Pro:out_test lindalan$ ls cellAAA Lindas-MacBook-Pro:out_test lindalan$ cd ..ls -bash: cd: ..ls: No such file or directory Lindas-MacBook-Pro:out_test lindalan$ cd .. Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd expected_summary/ Lindas-MacBook-Pro:expected_summary lindalan$ ls BCR_summary.txt clonotype_network_without_identifiers.pdf igblast_input_H.fa IMGT_gapped_db.tab clonotype_sizes.pdf igblast_input_L.fa changeo_input_H.tab clonotype_sizes.txt isotype_distribution.pdf changeo_input_H_clone-pass.tab concatenated_lineage_input.tab lineage_trees changeo_input_K.tab full_length_seqs.pdf reconstructed_lengths_BCR_H.pdf changeo_input_L.tab igblast_H.fmt7 reconstructed_lengths_BCR_H.txt changeo_input_L_clone-pass.tab igblast_H_db-modified.tab reconstructed_lengths_BCR_K.txt changeodb.tab igblast_H_db-modified_germ-pass.tab reconstructed_lengths_BCR_L.pdf clonotype_network_with_identifiers.dot igblast_L.fmt7 reconstructed_lengths_BCR_L.txt clonotype_network_with_identifiers.pdf igblast_L_db-modified.tab clonotype_network_without_identifiers.dot igblast_L_db-modified_germ-pass.tab Lindas-MacBook-Pro:expected_summary lindalan$ ls BCR_summary.txt clonotype_network_without_identifiers.pdf igblast_input_H.fa IMGT_gapped_db.tab clonotype_sizes.pdf igblast_input_L.fa changeo_input_H.tab clonotype_sizes.txt isotype_distribution.pdf changeo_input_H_clone-pass.tab concatenated_lineage_input.tab lineage_trees changeo_input_K.tab full_length_seqs.pdf reconstructed_lengths_BCR_H.pdf changeo_input_L.tab igblast_H.fmt7 reconstructed_lengths_BCR_H.txt changeo_input_L_clone-pass.tab igblast_H_db-modified.tab reconstructed_lengths_BCR_K.txt changeodb.tab igblast_H_db-modified_germ-pass.tab reconstructed_lengths_BCR_L.pdf clonotype_network_with_identifiers.dot igblast_L.fmt7 reconstructed_lengths_BCR_L.txt clonotype_network_with_identifiers.pdf igblast_L_db-modified.tab clonotype_network_without_identifiers.dot igblast_L_db-modified_germ-pass.tab Lindas-MacBook-Pro:expected_summary lindalan$ cd .. Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd out_test2 Lindas-MacBook-Pro:out_test2 lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 -g pdf /scratch/out_test2 usage: bracer [-h] [--ncores ] [--config_file
] [--resource_dir ] [--species SPECIES] [--loci LOCI [LOCI ...]] [--use_unfiltered] [--graph_format ] [--no_networks] [--IGH_networks] [--dist ] [--include_multiplets] [--infer_lineage] bracer: error: unrecognized arguments: -g /scratch/out_test2 Lindas-MacBook-Pro:out_test2 lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 --graph_format pdf /scratch/out_test2 Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in load_entry_point('bracer==0.1', 'console_scripts', 'bracer')() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch Task().run() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 712, in run subdirectories = next(os.walk(self.root_dir))[1] StopIteration Lindas-MacBook-Pro:out_test2 lindalan$ cd .. Lindas-MacBook-Pro:test_data lindalan$ ls cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results Lindas-MacBook-Pro:test_data lindalan$ cd out_test2 Lindas-MacBook-Pro:out_test2 lindalan$ ls cell1 cell2 cellA filtered_BCR_summary results Lindas-MacBook-Pro:out_test2 lindalan$ cd cellA Lindas-MacBook-Pro:cellA lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 --graph_format pdf /scratch/cellA usage: bracer [-h] [--ncores ] [--config_file
] [--resource_dir ] [--species SPECIES] [--loci LOCI [LOCI ...]] [--use_unfiltered] [--graph_format ] [--no_networks] [--IGH_networks] [--dist ] [--include_multiplets] [--infer_lineage] bracer: error: argument --graph_format/-f: expected one argument Lindas-MacBook-Pro:cellA lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 -f pdf /scratch/cellA Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in load_entry_point('bracer==0.1', 'console_scripts', 'bracer')() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch Task().run() File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 712, in run subdirectories = next(os.walk(self.root_dir))[1] StopIteration
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Linda-Lan
idalind
Hi bracer team,
I set up bracer through downloading the docker image directly, but ran error with test data as following. Do you have any suggestions for it?
Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer assemble -p 4 cell1 /scratch/out_test2 /scratch/cell1_1.fastq /scratch/cell1_2.fastq
Trimming raw reads
Detecting installed version of Cutadapt: 1.14 Trimming completed
Finding recombinant-derived reads
Attempting new assembly for ['BCR_H', 'BCR_K', 'BCR_L']
Detected average R1 read length: 50.0 Short read length detected. BraCeR will run two rounds of alignment for heavy chain
BCR_H
56433 reads; of these: 56433 (100.00%) were paired; of these: 34987 (62.00%) aligned concordantly 0 times 21446 (38.00%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
38.60% overall alignment rate 56433 reads; of these: 56433 (100.00%) were paired; of these: 55425 (98.21%) aligned concordantly 0 times 1008 (1.79%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
38.59% overall alignment rate
BCR_K
56433 reads; of these: 56433 (100.00%) were paired; of these: 48166 (85.35%) aligned concordantly 0 times 8267 (14.65%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
15.24% overall alignment rate
BCR_L
56433 reads; of these: 56433 (100.00%) were paired; of these: 31447 (55.72%) aligned concordantly 0 times 24986 (44.28%) aligned concordantly exactly 1 time 0 (0.00%) aligned concordantly >1 times
45.35% overall alignment rate
Assembling Trinity Contigs
BCR_H
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases
Wednesday, August 8, 2018: 23:45:43 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:45:44 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:45:44 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H Wednesday, August 8, 2018: 23:45:44 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:45:44 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:45:44 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_H_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:45:44 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:45:44 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa Wednesday, August 8, 2018: 23:45:44 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf
--------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.tmp
Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:45:46 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/inchworm.K25.L25.DS.fa.finished
-------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H/both.fa
------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:45:49 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 3 succeeded(3) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:45:57 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_H.Trinity.fasta ###################################################################
BCR_K
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases
Wednesday, August 8, 2018: 23:45:58 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:45:58 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:45:58 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K Wednesday, August 8, 2018: 23:45:58 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:45:58 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:45:58 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_K_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:45:58 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:45:58 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa Wednesday, August 8, 2018: 23:45:58 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf
--------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.tmp
Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:45:59 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/inchworm.K25.L25.DS.fa.finished
-------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K/both.fa
------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:46:02 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 1 succeeded(1) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:46:06 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_K.Trinity.fasta ###################################################################
BCR_L
Left read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_1.fastq' ]; Right read files: $VAR1 = [ '/scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_2.fastq' ]; Trinity version: Trinity-v2.4.0 ** NOTE: Latest version of Trinity is Trinity-v2.7.0-PRERELEASE, and can be obtained at: https://github.com/trinityrnaseq/trinityrnaseq/releases
Wednesday, August 8, 2018: 23:46:07 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 0 Wednesday, August 8, 2018: 23:46:07 CMD: java -Xmx64m -XX:ParallelGCThreads=2 -jar /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/ExitTester.jar 1 Wednesday, August 8, 2018: 23:46:07 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L Wednesday, August 8, 2018: 23:46:07 CMD: mkdir -p /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/chrysalis
-------------- Trinity Phase 1: Clustering of RNA-Seq Reads ---------------------
Converting input files. (in parallel)Wednesday, August 8, 2018: 23:46:07 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_1.fastq | seqtk-trinity seq -A - >> left.fa Wednesday, August 8, 2018: 23:46:07 CMD: cat /scratch/out_test2/cell1/aligned_reads/cell1_BCR_L_2.fastq | seqtk-trinity seq -A - >> right.fa Wednesday, August 8, 2018: 23:46:07 CMD: touch right.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: touch left.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: touch left.fa.ok right.fa.ok Wednesday, August 8, 2018: 23:46:07 CMD: cat left.fa right.fa > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa Wednesday, August 8, 2018: 23:46:07 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa.ok
----------- Jellyfish -------------------- -- (building a k-mer catalog from reads) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish count -t 4 -m 25 -s 100000000 --canonical /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish dump -L 1 mer_counts.jf > jellyfish.kmers.fa
Running CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/jellyfish/bin/jellyfish histo -t 4 -o jellyfish.kmers.fa.histo mer_counts.jf
--------------- Inchworm --------------------- -- (Linear contig construction from k-mers) --
Running CMD: /trinityrnaseq-Trinity-v2.4.0/Inchworm/bin//inchworm --kmers jellyfish.kmers.fa --run_inchworm -K 25 -L 25 --monitor 1 --DS --num_threads 4 --PARALLEL_IWORM > /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.tmp
Running CMD: mv /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.tmp /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa Wednesday, August 8, 2018: 23:46:09 CMD: touch /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/inchworm.K25.L25.DS.fa.finished
-------------------- Chrysalis ------------------------- -- (Contig Clustering & de Bruijn Graph Construction) --
inchworm_target: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa bowite_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa chrysalis_reads_fa: /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L/both.fa
------------ Trinity Phase 2: Assembling Clusters of Reads ---------------------
Wednesday, August 8, 2018: 23:46:13 CMD: /trinityrnaseq-Trinity-v2.4.0/trinity-plugins/parafly/bin/ParaFly -c recursive_trinity.cmds -CPU 4 -v Number of Commands: 1 succeeded(1) 100% completed.
All commands completed successfully. :-)
** Harvesting all assembled transcripts into a single multi-fasta file...
Wednesday, August 8, 2018: 23:46:21 CMD: find read_partitions/ -name '*inity.fasta' | /trinityrnaseq-Trinity-v2.4.0/util/support_scripts/partitioned_trinity_aggregator.pl TRINITY_DN > Trinity.fasta.tmp
################################################################### Butterfly assemblies are written to /scratch/out_test2/cell1/Trinity_output/Trinity_cell1_BCR_L.Trinity.fasta ###################################################################
Running BLAST
Performing Blast on ['BCR_H', 'BCR_K', 'BCR_L']
BCR_H
BCR_K
BCR_L
Running IgBLAST
Ig_seqtype: Ig Performing IgBlast on ['BCR_H', 'BCR_K', 'BCR_L']
BCR_H
BCR_K
BCR_L
ALIGNER_OUTPUT> cell1_BCR_H.fmt7 SEQ_FILE> cell1_BCR_H.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (3) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_H_db-pass.tab PASS> 1 FAIL> 2 END> MakeDb
ALIGNER_OUTPUT> cell1_BCR_K.fmt7 SEQ_FILE> cell1_BCR_K.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (1) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_K_db-pass.tab PASS> 1 FAIL> 0 END> MakeDb
ALIGNER_OUTPUT> cell1_BCR_L.fmt7 SEQ_FILE> cell1_BCR_L.Trinity.fasta NO_PARSE> False PARTIAL> False SCORES> True REGIONS> True
PROGRESS> 23:46:34 [Done ] 0.0 min
PROGRESS> 23:46:34 [####################] 100% (1) 0.0 min
OUTPUT> /scratch/out_test2/cell1/IgBLAST_output/cell1_BCR_L_db-pass.tab PASS> 1 FAIL> 0 END> MakeDb
Running Kallisto
Making Kallisto indices
[build] loading fasta file /scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.fa [build] k-mer length: 31 [build] warning: clipped off poly-A tail (longer than 10) from 1549 target sequences [build] warning: replaced 4 non-ACGUT characters in the input sequence with pseudorandom nucleotides [build] counting k-mers ... Traceback (most recent call last): File "/usr/local/bin/bracer", line 11, in
load_entry_point('bracer==0.1', 'console_scripts', 'bracer')()
File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/launcher.py", line 43, in launch
Task().run()
File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 369, in run
self.quantify(cell)
File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/tasks.py", line 605, in quantify
self.trimmed_fastq2, self.keep_trimmed_reads)
File "/usr/local/lib/python3.5/dist-packages/bracer-0.1-py3.5.egg/bracerlib/bracer_func.py", line 2110, in quantify_with_kallisto
subprocess.check_call(index_command)
File "/usr/lib/python3.5/subprocess.py", line 271, in check_call
raise CalledProcessError(retcode, cmd)
subprocess.CalledProcessError: Command '['/kallisto_linux-v0.43.1/kallisto', 'index', '-i', '/scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.idx', '/scratch/out_test2/cell1/expression_quantification/kallisto_index/cell1_transcriptome.fa']' returned non-zero exit status -9
Lindas-MacBook-Pro:test_data lindalan$ ls
cell1_1.fastq cell1_2.fastq expected_summary out_test out_test2 results
Lindas-MacBook-Pro:test_data lindalan$ cd out_test
Lindas-MacBook-Pro:out_test lindalan$ ls
cellAAA
Lindas-MacBook-Pro:out_test lindalan$ cd cellAAA/
Lindas-MacBook-Pro:cellAAA lindalan$ ls
BLAST_output Trinity_output expression_quantification trimmed_reads
IgBLAST_output aligned_reads filtered_BCR_seqs unfiltered_BCR_seqs
Lindas-MacBook-Pro:cellAAA lindalan$ cd /Users/lindalan/docker/bracer/bracer/test_data
Lindas-MacBook-Pro:test_data lindalan$ docker run -it --rm -v $PWD:/scratch -w /scratch teichlab/bracer summarise -p 4 -g pdf /scratch/out_test2
usage: bracer [-h] [--ncores ] [--config_file ]
[--resource_dir ] [--species SPECIES]
[--loci LOCI [LOCI ...]] [--use_unfiltered]
[--graph_format ] [--no_networks] [--IGH_networks]
[--dist ] [--include_multiplets] [--infer_lineage]