Mapping Rate: 0%

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Carlos Guzman

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Aug 17, 2016, 12:23:37 PM8/17/16

to Sailfish Users Group

I'm trying out Salmon and having a surprisingly difficult time getting it to work.

Here is the latest error:

salmon --no-version-check quant -i '/dataA/data/hg38_ref/salmon_transcripts.idx' -l MU -1 /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R1_001_val_1.fq.gz -2 /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R2_001_val_2.fq.gz -p 30 -o '/dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/salmon_quant' 
# salmon (mapping-based) v0.6.0
# [ program ] => salmon 
# [ command ] => quant 
# [ index ] => { /dataA/data/hg38_ref/salmon_transcripts.idx }
# [ libType ] => { MU }
# [ mates1 ] => { /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R1_001_val_1.fq.gz }
# [ mates2 ] => { /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R2_001_val_2.fq.gz }
# [ threads ] => { 30 }
# [ output ] => { /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/salmon_quant }
Logs will be written to /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/salmon_quant/logs
there is 1[2016-08-17 11:21:17.819] [jointLog] [info] parsing read library format
 lib
Loading 32-bit quasi indextcmalloc: large alloc 1125023744 bytes == 0x1f07a000 @  0x50c9c2 0x7f95b5 0x614d3a 0x619fce 0x5abf05 0x5bc466 0x5bd038 0x5817ba 0x50ea10 0x7ffb714aff45 0x51593d
[2016-08-17 11:21:17.870] [jointLog] [info] Loading Quasi index
tcmalloc: large alloc 4294967296 bytes == 0x62162000 @  0x50c9c2 0x7f9433 0x615803 0x615e1c 0x612cfe 0x612f8e 0x7ffb73139a80 0x61a641 0x7e8d20 0x7ffb73134184 0x7ffb7158837d
[2016-08-17 11:21:17.870] [stderrLog] [info] Loading Suffix Array 
[2016-08-17 11:21:17.870] [stderrLog] [info] Loading Position Hash
[2016-08-17 11:21:18.415] [stderrLog] [info] Loading Transcript Info 
[2016-08-17 11:21:18.583] [stderrLog] [info] Loading Rank-Select Bit Array
[2016-08-17 11:21:18.663] [stderrLog] [info] There were 173259 set bits in the bit array
[2016-08-17 11:21:18.680] [stderrLog] [info] Computing transcript lengths
[2016-08-17 11:21:18.681] [stderrLog] [info] Waiting to finish loading hash
Index contained 173259 targets
[2016-08-17 11:21:22.744] [jointLog] [info] done
[2016-08-17 11:21:22.744] [stderrLog] [info] Done loading index












[2016-08-17 11:21:22.975] [jointLog] [info] Computed 0 rich equivalence classes for further processing
[2016-08-17 11:21:22.975] [jointLog] [info] Counted 0 total reads in the equivalence classes 
[2016-08-17 11:21:32.288] [jointLog] [warning] Only 0 fragments were mapped, but the number of burn-in fragments was set to 5000000.
The effective lengths have been computed using the observed mappings.

[2016-08-17 11:21:32.288] [jointLog] [warning] Something seems to be wrong with the calculation of the mapping rate.  The recorded ratio is likely wrong.  Please file this as a bug report.

[2016-08-17 11:21:32.288] [jointLog] [info] Mapping rate = 0%

[2016-08-17 11:21:32.288] [jointLog] [info] finished quantifyLibrary()
[2016-08-17 11:21:32.288] [jointLog] [info] Starting optimizer
[2016-08-17 11:21:32.325] [jointLog] [info] Marked 0 weighted equivalence classes as degenerate
[2016-08-17 11:21:32.326] [jointLog] [info] iteration = 0 | max rel diff. = -1.79769e+308
[2016-08-17 11:21:32.346] [jointLog] [info] iteration = 50 | max rel diff. = -1.79769e+308
[2016-08-17 11:21:32.359] [jointLog] [info] Finished optimizer
[2016-08-17 11:21:32.359] [jointLog] [info] writing output

I've tried other library strand combinations, but they all end up with the same error.

Here's the infer_experiment.py output from RSeqC:

This is PairEnd Data
Fraction of reads failed to determine: 0.0375
Fraction of reads explained by "1++,1--,2+-,2-+": 0.0261
Fraction of reads explained by "1+-,1-+,2++,2--": 0.9365

Rob

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Aug 17, 2016, 4:28:05 PM8/17/16

to Sailfish Users Group

Hi Carlos,

Reading directly from gzipped input is only supported in v0.7.0. It looks like you're running v0.6.0, so you'll have to use process substitution to produce uncompressed reads. Try the following command:

salmon --no-version-check quant -i /dataA/data/hg38_ref/salmon_transcripts.idx -l IU -1 <(gunzip -c /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R1_001_val_1.fq.gz) -2 <(gunzip -c /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/trimgalore/Ctrl_1_S1_R2_001_val_2.fq.gz) -p 30 -o /dataA/data/Curtis_RNAseq_08-15-2016/_/2016_08_05_CPL160_1400_0/raw/Sample_Ctrl_1/salmon_quant

Note the `'` surrounding the input arguments aren't required. Could you let me know if this works for you?

--Rob

Carlos Guzman

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Aug 17, 2016, 5:16:02 PM8/17/16

to Sailfish Users Group

That seems to have solved the problem. Much appreciated Rob!

Napoleon Little

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Jan 10, 2017, 11:40:43 PM1/10/17

to Sailfish Users Group

I have the same problem,Mapping rate : 0%, codes are as follows,

expList<-cn_salmon(stQuery, refDir="ref/",salmonIndex=iFileMouse, salmonPath=pathToSalmon)
determining read length
Trimming reads
When using programs that use GNU Parallel to process data for publication please cite:

O. Tange (2011): GNU Parallel - The Command-Line Power Tool,
;login: The USENIX Magazine, February 2011:42-47.

This helps funding further development; and it won't cost you a cent.
Or you can get GNU Parallel without this requirement by paying 10000 EUR.

To silence this citation notice run 'parallel --bibtex' once or use '--no-notice'.

This is cutadapt 1.9.1 with Python 2.7.12
Command line parameters: -m 30 -u 84 -u -83 -o ./SRR2042996_trimmed.fq SRR2042996.fastq
Trimming 0 adapters with at most 10.0% errors in single-end mode ...
Finished in 52.64 s (4 us/read; 13.64 M reads/minute).

=== Summary ===

Total reads processed:              11,963,935
Reads with adapters:                         0 (0.0%)
Reads that were too short:          11,894,824 (99.4%)
Reads written (passing filters):        69,111 (0.6%)

Total basepairs processed: 1,205,200,973 bp
Total written (filtered):      2,867,074 bp (0.2%)

This is cutadapt 1.9.1 with Python 2.7.12
Command line parameters: -m 30 -u 18 -u -17 -o ./SRR2042995_trimmed.fq SRR2042995.fastq
Trimming 0 adapters with at most 10.0% errors in single-end mode ...
Finished in 90.44 s (7 us/read; 8.39 M reads/minute).

=== Summary ===

Total reads processed:              12,645,225
Reads with adapters:                         0 (0.0%)
Reads that were too short:           2,120,789 (16.8%)
Reads written (passing filters):    10,524,436 (83.2%)

Total basepairs processed: 1,322,516,359 bp
Total written (filtered):    859,655,287 bp (65.0%)

This is cutadapt 1.9.1 with Python 2.7.12
Command line parameters: -m 30 -u 32 -u -32 -o ./SRR2042994_trimmed.fq SRR2042994.fastq
Trimming 0 adapters with at most 10.0% errors in single-end mode ...
Finished in 89.29 s (7 us/read; 8.67 M reads/minute).

=== Summary ===

Total reads processed:              12,900,445
Reads with adapters:                         0 (0.0%)
Reads that were too short:           3,154,202 (24.5%)
Reads written (passing filters):     9,746,243 (75.5%)

Total basepairs processed: 1,516,798,037 bp
Total written (filtered):    680,462,683 bp (44.9%)

Salmon
When using programs that use GNU Parallel to process data for publication please cite:

O. Tange (2011): GNU Parallel - The Command-Line Power Tool,
;login: The USENIX Magazine, February 2011:42-47.

This helps funding further development; and it won't cost you a cent.
Or you can get GNU Parallel without this requirement by paying 10000 EUR.

To silence this citation notice run 'parallel --bibtex' once or use '--no-notice'.

[2017-01-09 12:36:34.261] [jointLog] [info] parsing read library format
[2017-01-09 12:36:34.261] [jointLog] [info] There is 1 library.
[2017-01-09 12:36:34.416] [jointLog] [info] Loading Quasi index
[2017-01-09 12:36:34.438] [jointLog] [info] Loading 32-bit quasi index
[2017-01-09 12:36:34.438] [stderrLog] [info] Loading Suffix Array
[2017-01-09 12:36:34.468] [stderrLog] [info] Successfully loaded position hash
[2017-01-09 12:36:41.398] [stderrLog] [info] Loading Transcript Info
[2017-01-09 12:36:43.974] [stderrLog] [info] Loading Rank-Select Bit Array
[2017-01-09 12:36:44.383] [stderrLog] [info] There were 106125 set bits in the bit array
[2017-01-09 12:36:44.425] [stderrLog] [info] Computing transcript lengths
[2017-01-09 12:36:44.425] [stderrLog] [info] Waiting to finish loading hash
[2017-01-09 12:36:44.485] [stderrLog] [info] Done loading index
[2017-01-09 12:36:44.492] [jointLog] [info] done
[2017-01-09 12:36:44.492] [jointLog] [info] Index contained 106125 targets
[2017-01-09 12:36:45.816] [jointLog] [info] Computed 0 rich equivalence classes for further processing
[2017-01-09 12:36:45.816] [jointLog] [info] Counted 0 total reads in the equivalence classes
[2017-01-09 12:36:45.852] [jointLog] [warning] Only 0 fragments were mapped, but the number of burn-in fragments was set to 5000000.

The effective lengths have been computed using the observed mappings.

[2017-01-09 12:36:45.852] [jointLog] [info] Mapping rate = 0%

[2017-01-09 12:36:45.914] [jointLog] [info] finished quantifyLibrary()
[2017-01-09 12:36:45.954] [jointLog] [info] Starting optimizer
[2017-01-09 12:36:46.302] [jointLog] [info] Marked 0 weighted equivalence classes as degenerate
[2017-01-09 12:36:46.303] [jointLog] [info] iteration = 0 | max rel diff. = 0.510182
[2017-01-09 12:36:46.477] [jointLog] [info] iteration = 100 | max rel diff. = -1.79769e+308
[2017-01-09 12:36:46.491] [jointLog] [info] Finished optimizer
[2017-01-09 12:36:46.491] [jointLog] [info] writing output

Version Info: This is the most recent version of Salmon.
### salmon (mapping-based) v0.7.2
### [ program ] => salmon
### [ command ] => quant
### [ threads ] => { 5 }
### [ index ] => { ref//salmon.index.mouse.122116 }
### [ libType ] => { U }
### [ unmatedReads ] => { SRR2042996_trimmed.fq }
### [ output ] => { salmonRes_SRR2042996 }
Logs will be written to salmonRes_SRR2042996/logs

[2017-01-09 12:36:39.470] [jointLog] [info] parsing read library format
[2017-01-09 12:36:39.470] [jointLog] [info] There is 1 library.
[2017-01-09 12:36:39.606] [jointLog] [info] Loading Quasi index
[2017-01-09 12:36:39.606] [jointLog] [info] Loading 32-bit quasi index
[2017-01-09 12:36:39.606] [stderrLog] [info] Loading Suffix Array
[2017-01-09 12:36:39.608] [stderrLog] [info] Successfully loaded position hash
[2017-01-09 12:36:41.398] [stderrLog] [info] Loading Transcript Info
[2017-01-09 12:36:43.974] [stderrLog] [info] Loading Rank-Select Bit Array
[2017-01-09 12:36:44.383] [stderrLog] [info] There were 106125 set bits in the bit array
[2017-01-09 12:36:44.424] [stderrLog] [info] Computing transcript lengths
[2017-01-09 12:36:44.424] [stderrLog] [info] Waiting to finish loading hash
[2017-01-09 12:36:44.485] [stderrLog] [info] Done loading index
[2017-01-09 12:36:44.492] [jointLog] [info] done
[2017-01-09 12:36:44.492] [jointLog] [info] Index contained 106125 targets
[2017-01-09 12:38:57.005] [jointLog] [info] Computed 0 rich equivalence classes for further processing
[2017-01-09 12:38:57.005] [jointLog] [info] Counted 0 total reads in the equivalence classes
[2017-01-09 12:38:57.029] [jointLog] [warning] Only 0 fragments were mapped, but the number of burn-in fragments was set to 5000000.

The effective lengths have been computed using the observed mappings.

[2017-01-09 12:38:57.029] [jointLog] [info] Mapping rate = 0%

[2017-01-09 12:38:57.029] [jointLog] [info] finished quantifyLibrary()
[2017-01-09 12:38:57.043] [jointLog] [info] Starting optimizer
[2017-01-09 12:38:57.328] [jointLog] [info] Marked 0 weighted equivalence classes as degenerate
[2017-01-09 12:38:57.329] [jointLog] [info] iteration = 0 | max rel diff. = 0.510182
[2017-01-09 12:38:57.404] [jointLog] [info] iteration = 100 | max rel diff. = -1.79769e+308
[2017-01-09 12:38:57.406] [jointLog] [info] Finished optimizer
[2017-01-09 12:38:57.406] [jointLog] [info] writing output

Version Info: This is the most recent version of Salmon.
### salmon (mapping-based) v0.7.2
### [ program ] => salmon
### [ command ] => quant
### [ threads ] => { 5 }
### [ index ] => { ref//salmon.index.mouse.122116 }
### [ libType ] => { U }
### [ unmatedReads ] => { SRR2042994_trimmed.fq }
### [ output ] => { salmonRes_SRR2042994 }
Logs will be written to salmonRes_SRR2042994/logs

processed 9500000 fragments
hits: 0; hits per frag: 0

[2017-01-09 12:36:38.986] [jointLog] [info] parsing read library format
[2017-01-09 12:36:38.986] [jointLog] [info] There is 1 library.
[2017-01-09 12:36:39.126] [jointLog] [info] Loading Quasi index
[2017-01-09 12:36:39.126] [jointLog] [info] Loading 32-bit quasi index
[2017-01-09 12:36:39.126] [stderrLog] [info] Loading Suffix Array
[2017-01-09 12:36:39.128] [stderrLog] [info] Successfully loaded position hash
[2017-01-09 12:36:41.398] [stderrLog] [info] Loading Transcript Info
[2017-01-09 12:36:43.974] [stderrLog] [info] Loading Rank-Select Bit Array
[2017-01-09 12:36:44.383] [stderrLog] [info] There were 106125 set bits in the bit array
[2017-01-09 12:36:44.427] [stderrLog] [info] Computing transcript lengths
[2017-01-09 12:36:44.427] [stderrLog] [info] Waiting to finish loading hash
[2017-01-09 12:36:44.485] [stderrLog] [info] Done loading index
[2017-01-09 12:36:44.492] [jointLog] [info] done
[2017-01-09 12:36:44.492] [jointLog] [info] Index contained 106125 targets
[2017-01-09 12:39:25.050] [jointLog] [info] Computed 0 rich equivalence classes for further processing
[2017-01-09 12:39:25.050] [jointLog] [info] Counted 0 total reads in the equivalence classes
[2017-01-09 12:39:25.063] [jointLog] [warning] Only 0 fragments were mapped, but the number of burn-in fragments was set to 5000000.

The effective lengths have been computed using the observed mappings.

[2017-01-09 12:39:25.063] [jointLog] [info] Mapping rate = 0%

[2017-01-09 12:39:25.063] [jointLog] [info] finished quantifyLibrary()
[2017-01-09 12:39:25.064] [jointLog] [info] Starting optimizer
[2017-01-09 12:39:25.098] [jointLog] [info] Marked 0 weighted equivalence classes as degenerate
[2017-01-09 12:39:25.099] [jointLog] [info] iteration = 0 | max rel diff. = 0.510182
[2017-01-09 12:39:25.132] [jointLog] [info] iteration = 100 | max rel diff. = -1.79769e+308
[2017-01-09 12:39:25.134] [jointLog] [info] Finished optimizer
[2017-01-09 12:39:25.134] [jointLog] [info] writing output

Version Info: This is the most recent version of Salmon.
### salmon (mapping-based) v0.7.2
### [ program ] => salmon
### [ command ] => quant
### [ threads ] => { 5 }
### [ index ] => { ref//salmon.index.mouse.122116 }
### [ libType ] => { U }
### [ unmatedReads ] => { SRR2042995_trimmed.fq }
### [ output ] => { salmonRes_SRR2042995 }
Logs will be written to salmonRes_SRR2042995/logs

processed 10500000 fragments
hits: 0; hits per frag: 0

./salmonRes_SRR2042994/quant.sf
./salmonRes_SRR2042995/quant.sf
./salmonRes_SRR2042996/quant.sf