Scasa:v1.0.1 docker shows error

[morgan-th]

I am trying to use the scasav1.0.1 docker in a Mac M1 system and I get the following error when running the test dataset. The main issue is that it says that it cannot read the input file right before "Starting the Pufferfish indexing by reading the GFA binary file." line.

I have already changed the "/path/to" to the scasa path in the docker_params.sh file and changed the -v argument of the if statements in the runScasaDocker.sh file to the -n argument.

The entire console log is shown below:

---

                 You are running Scasa v1.0.1 using docker ....

---

Loading parameters from file...
WARNING: The requested image's platform (linux/amd64) does not match the detected host platform (linux/arm64/v8) and no specific platform was requested

##############################################################

SCASA V1.0.1

SINGLE CELL TRANSCRIPT QUANTIFICATION TOOL

Version Date: 2022-03-24

FOR ANY ISSUES, CONTACT: [email protected]

https://github.com/eudoraleer/scasa/

##############################################################

Directory /source/output already exists. Writing into existing directory..
mkdir: cannot create directory '/source/output/SCASA_My_Project_20230322024150/': File exists

Preparing for alignment..
Indexing reference..
Directory /source/output/SCASA_My_Project_20230322024150/0PRESETS//REF_INDEX/ already exists. Writing into existing directory..
Version Info: ### PLEASE UPGRADE SALMON ###

A newer version of salmon with important bug fixes and improvements is available.

The newest version, available at https://github.com/COMBINE-lab/salmon/releases
contains new features, improvements, and bug fixes; please upgrade at your
earliest convenience.

Sign up for the salmon mailing list to hear about new versions, features and updates at:
https://oceangenomics.com/subscribe
###[2023-03-22 02:41:50.723] [jLog] [warning] The salmon index is being built without any decoy sequences. It is recommended that decoy sequence (either computed auxiliary decoy sequence or the genome of the organism) be provided during indexing. Further details can be found at https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode.
[2023-03-22 02:41:50.725] [jLog] [info] building index
out : /source/output/SCASA_My_Project_20230322024150/0PRESETS//REF_INDEX/
[2023-03-22 02:41:50.730] [puff::index::jointLog] [info] Running fixFasta

[Step 1 of 4] : counting k-mers

[2023-03-22 02:42:49.745] [puff::index::jointLog] [warning] Removed 237 transcripts that were sequence duplicates of indexed transcripts.
[2023-03-22 02:42:49.746] [puff::index::jointLog] [warning] If you wish to retain duplicate transcripts, please use the --keepDuplicates flag
[2023-03-22 02:42:49.751] [puff::index::jointLog] [info] Replaced 5 non-ATCG nucleotides
[2023-03-22 02:42:49.751] [puff::index::jointLog] [info] Clipped poly-A tails from 12501 transcripts
wrote 70629 cleaned references
[2023-03-22 02:43:20.214] [puff::index::jointLog] [info] Filter size not provided; estimating from number of distinct k-mers
[2023-03-22 02:43:29.656] [puff::index::jointLog] [info] ntHll estimated 84081876 distinct k-mers, setting filter size to 2^31
Threads = 2
Vertex length = 31
Hash functions = 5
Filter size = 2147483648
Capacity = 2
Files:
/source/output/SCASA_My_Project_20230322024150/0PRESETS//REF_INDEX/ref_k31_fixed.fa

Round 0, 0:2147483648
Pass Filling Filtering
1 126 216
2 41 1
True junctions count = 266516
False junctions count = 405083
Hash table size = 671599
Candidate marks count = 4093845

Error: Can't read from a temporary file
allowedIn: 12
error: Can't read the input file
[2023-03-22 02:50:25.299] [puff::index::jointLog] [info] Starting the Pufferfish indexing by reading the GFA binary file.
[2023-03-22 02:50:25.300] [puff::index::jointLog] [info] Setting the index/BinaryGfa directory /source/output/SCASA_My_Project_20230322024150/0PRESETS//REF_INDEX
size = 0

| Loading contigs | Time = 732.71 us

size = 0

| Loading contig boundaries | Time = 794.58 us

Number of ones: 0
Number of ones per inventory item: 512
Inventory entries filled: 1
274911049145
[2023-03-22 02:50:25.303] [puff::index::jointLog] [info] Done wrapping the rank vector with a rank9sel structure.
Finnished indexing reference..
Begins pseudo-alignment..
nohup: redirecting stderr to stdout
Congratulations! Pseudo-alignment has completed in 30 seconds!
Scasa quantification has started..
Begin Scasa quantification for sample Sample_01_S1_L001..
Error in file(con, "r") : cannot open the connection
Calls: readLines -> file
In addition: Warning message:
In file(con, "r") :
cannot open file '/source/output/SCASA_My_Project_20230322024150/1ALIGN//Sample_01_S1_L001_alignout/alevin/bfh.txt': No such file or directory
Execution halted
Loading required package: iterators
Loading required package: parallel
Error in readChar(con, 5L, useBytes = TRUE) : cannot open the connection
Calls: load -> readChar
In addition: Warning message:
In readChar(con, 5L, useBytes = TRUE) :
cannot open compressed file '/source/output/SCASA_My_Project_20230322024150/2QUANT/Sample_01_S1_L001_quant/Sample_eqClass.RData', probable reason 'No such file or directory'
Execution halted
Error in readChar(con, 5L, useBytes = TRUE) : cannot open the connection
Calls: load -> readChar
In addition: Warning message:
In readChar(con, 5L, useBytes = TRUE) :
cannot open compressed file '/source/output/SCASA_My_Project_20230322024150/2QUANT//Sample_01_S1_L001_quant//scasa_isoform_expression.RData', probable reason 'No such file or directory'
Execution halted

[nghiavtr]

Hi @morgan-th

Thank you for using Scasa in your research.

For the issue, we usually test and run Scasa docker in linux environment such as ubuntu, so we are not sure about possible issues occurring in Mac os. Furthermore, you mentioned that
"changed the -v argument of the if statements in the runScasaDocker.sh file to the -n argument."
Could you let me know why you made the change? Normally we dont need to revise the code in runScasaDocker.sh file, but change the proper values of the parameters in docker_params.sh file

Best,
Nghia

[morgan-th]

Thank you @nghiavtr for you reply!

The -v argument was giving me a syntax error in the if statement lines and revising it to the -n argument solved this issue.

[nghiavtr]

In bash, -v is used to check whether a variable is set or not. I am not sure which similar option is used in the terminal of Mac OS.
Anyway, I guess the issue was possibly due to different operation systems, so I suggest you running Scasa in a linux environment. e.g, ubuntu.

Best,
Nghia