MinPipe

MinPipe is a minimal but fully logged pipeline or workflow to be used for a series of RNA-seq data analysis from raw data to differentially expressed genes by using a mixture of Python, R and Shell script mainly for workspace management.

Installing

Rewritten version of installers include:

• env.yml: The yaml file that sets specification for environment building used in miniconda/mamba. It lists all needed packages to run every part of MinPipe.
• installers/conda.sh: A shell script to download and install miniconda, mamba and create the needed environment. Go to the project root directory and run bash installers/conda.sh.

Using the main pipeline

• Run python3 minpipe.py -h or python3 minpipe.py --help for a help message
• Example of paired-ended analysis without pre-built index:
  • python3 minpipe.py -c _read1 _read2 -s AT1_G1 AT2_G1 AT1_G2 AT2_G2 -t MMU_39.fastq.gz --threads 4 -b 100
• Example of single-ended analysis with pre-built index:
  • python3 minpipe.py -s AT1_G1 AT2_G1 AT1_G2 AT2_G2 -i MMU_39.idx --threads 4 -b 100

Arguments

• -c or --complement is the complement for paired-ended file names, if read 1 is always sample_R1.fq.gz and read 2 is sample_R2.fq.gz use -c _R1 _R2 or --complement _R1 _R2 so the code will iterate over samples with this complementary name.
• -s or --samples is the list of samples used to integrate with complement and iterate in the directory, e.g. -s sample1 sample2 sample3 or --sample sample1 sample2 sample3 the program will iterate as sample1_R1.fq.gz and sample1_R2.fq.gz as paired-ended.
• -i or --index is the Name of the index file to be used in pseudoalignment. Either index or transcript has to be passed.
• -t or --transcript is the Name of the transcript file to be indexed. mmu or hsa can be passed so the transcript will be downloaded automatically and index will be built.
• --threads refers to the number of threads to be used in quantification for Kallisto. Default: 1.
• -b or --bootstrap is the number of bootstrap samples. Default: 100
• --single is the flag to indicate single-ended quantification without complements. An optional argument.
• --ext-qc is a flag to indicate that will have extensive QC. MAY NEED MORE FILES
• --json pass the Json file name that has to be located inside the input folder. The user can create separated folders inside the input, e.g. input/params/parameters.json.
• --yaml pass the YAML/YML file name that has to be located inside the input folder. The user can do the same as the Json file creating folders, e.g. input/params/parameters.yml.

How to work with Kallisto results using Sleuth R package

• Run Rscript minpipe.R [arguments]
• Pass Rscript minpipe.R -h to see the help text

Arguments

• -f or --file is the argument needed for the metadata.txt, passing as a path/to/metadata.txt
• -o or --organism is the name of the organism to be used for gene annotation, either mmu or hsa, others will be supported lately
• -p or --path is the path/to/kallisto/results where it would have SAMPLE_FOLDER/abundance.h5 files.
• -r or --results is the name of the path/to/save/results for tables and visualization.
• -s or --separator string used as a separator for metadata file. Default is ;
• --no-volcano is a flag that will force no volcano image creation