evatool 0.1.9

Extracellular Vesicles small RNAs Abundance and quantification tool

EVAtool

---

Background

non-coding RNAs (ncRNA) in extracellular vesicles (EVs) role as important agents of cell-to-cell communication. The abundance of transcripts varies in different states, especially in the tumors. Therefore, quantification of ncRNAs in EVs plays an important role in the process of exploring cancer diagnostic biomarkers.

Requires

python >= 3.5

Softwares

samtools = 1.12
bowtie2 = 2.4.2
fastq-dump = 2.10.9
trimmomatic-0.39.jar
bedtools = 2.30.0

Usage

• Reference and config file download

  • wget "http://bioinfo.life.hust.edu.cn/EVAtool/ref/refs.zip"

  • The details of Reference:

    • Homo_sapiens.GRCh38.86.chr.gtf
    • Homo_sapiens_v86
    • miRNA miRBase V22
    • snoRNA snoDB V1.2.1
    • tRNA GtRNAdb V18.1
    • piRNA RNAcentral V17
    • rRNA RNAcentral V17
    • snRNA NCBI GenBank
    • YRNA NCBI GenBank

  • The content of config file:

    • "trimmomatic_sRNA_para": "2:10:4:5 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:15",
    • "tag_cut": "0",
    • "cpu_number": "8",
    • "RPM": "total",
    • "bowtie_para_4_miRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 2 -L 15 -i S,1,1.15",
    • "bowtie_para_4_rRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 1 -L 12 -i S,1,1.15",
    • "bowtie_para_4_tRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 1 -L 12 -i S,1,1.15",
    • "bowtie_para_4_piRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 1 -L 12 -i S,1,1.15",
    • "bowtie_para_4_snRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 3 -L 10 -i S,1,1.15",
    • "bowtie_para_4_snoRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -L 12 -i S,1,1.15",
    • "bowtie_para_4_YRNA": "--no-head --no-sq -D 15 -R 2 -N 1 -k 1 -L 12 -i S,1,1.15",
    • "bowtie_para_4_genome": "-D 15 -R 2 -N 1 -k 1 -L 10 -i S,1,1.15",
    • "adp_path": "/refs/sRNA.fa",
    • "miRNA_index": "/refs/miRNA/hsa.hairpin.fa",
    • "mirbase": "/refs/miRNA/hsa.mirbase.txt",
    • "piRNA_index": "/refs/piRNA/piRNA.fa",
    • "YRNA_index": "/refs/YRNA/YRNA.fa",
    • "snoRNA_index": "/refs/snoRNA/snoRNA.fa",
    • "snRNA_index": "/refs/snRNA/snRNA.fa",
    • "rRNA_index": "/refs/rRNA/rRNA.fa",
    • "tRNA_index": "/refs/tRNA/tRNA.fa",
    • "genome_index": "/refs/Homo_sapiens_v86/Homo_sapiens_v86.chromosomes.fasta",
    • "gtf_annotation": "/refs/Homo_sapiens.GRCh38.86.chr.gtf",
    • "trans_bed_annotation": "/refs/four_elements.sort.bed",
    • "exon_bed_annotation": "/refs/two_elements.sort.bed"

• Install evatool

pip install evatool

• Example

>>> evatool -i [absolute path]/example.fastq.gz -o [output directory]

• Input parameters (Enter evatool -h for more details)

#-i: sra file with path (required)
#-o: output directory (required)
#-c: configure file with path ( not required)
#-n: ncRNA type list (not required)
#-h: More details about usages

• The other two ways when use EVAtool:

1. Download latest EVAtool docker image

# installation
docker pull guobioinfolab/evatool:latest
# run evatool
docker run -it --rm -v evatool -i [absolute path of the input data] -o [absolute path of the output directory]

2. pip install evatool

>>>from utils.fastq import Fastq
>>>from utils.config import Config
>>>from utils.logger import Logger
>>> def test_config():
...     from utils.config import Config
...     config = Config(configfile="./refs/reference_config.json")
...     print(config.config)

>>> test_config()

Modules in evatool

1. Logger
   • Produce a log file that records the events when the evatool is running.
   • Logger(logfile = Path)
2. Config
   • Tansfer the content of config file to each module.
   • Config(config = Path)
3. Fastq
   • Transform the raw .sra file to .fastq or .fastq.gz, quality control (QC) and adapter trimming.
   • Fastq(inputfile, outputdir, config=Config, log=Logger, ncrna_lst=ncrna_lst)
4. Tag
   • Statistics the distribution of various types of reads and transformed the trimmed file into .fasta.
   • Tag(fastq=Fastq(inputfile, outputdir, config=Config, log=Logger, ncrna_lst=ncrna_lst))
5. SAM
   • Map reads to human genome and other types ncRNA reference.
   • Tag(fastq=Fastq(inputfile, outputdir, config=Config, log=Logger, ncrna_lst=ncrna_lst))
6. Bam
   • Transform sam file to bam file.
   • Bam(fastq=Fastq, tag=Tag)
7. Stat
   • Quantification and RPM normalization.
   • Stat(fastq=Fastq, tag=Tag)
8. Plot
   • Visualize output results.
   • Plot(inputfile, outputdir)
9. Report
   • Provide online results report.
   • Report(inputfile, outputdir, config=Config, ncrna_lst=ncrna_lst))

Advanced usage

1. Use each module independently

After installing evatool, users only need to import the interested package(s) in the python interpreter to use the corresponding module.

e.g.

>>>from utils.fastq import Fastq
>>>from utils.config import Config
>>>from utils.logger import Logger
>>> def test_config():
...     from utils.config import Config
...     config = Config(configfile="./refs/reference_config.json")
...     print(config.config)

>>> test_config()

2. Custom ncRNA type(s)

   • Based on existing reference sequences

     • change the input parameter of ncRNA type list, the default ncRNAs are include 7 types: "miRNA" "rRNA" "tRNA" "piRNA" "snoRNA" "snRNA" "YRNA".

   • Add other type reference sequences

     • Three steps :
     1. Add the ncRNA reference and index into refs;

     mkdir [ncRNA name]
     add the reference and index to the directory
     

     2. Change the input parameter of ncRNA type list;

     -n  "miRNA" "rRNA" "tRNA"
     

     3. Add the ncRNA name and reference directory in config file as following the existing ways.

     "miRNA_index": "/refs/miRNA/hsa.hairpin.fa"
     

Directory tree (main)

├── bin
│   ├── bedtools
│   ├── bowtie2
│   ├── bowtie2-align-l
│   ├── bowtie2-align-l-debug
│   ├── bowtie2-align-s
│   ├── bowtie2-align-s-debug
│   ├── fastq-dump
│   ├── samtools
│   └── trimmomatic-0.39.jar
├── __init__.py
├── main.py
├── resource
│   ├── __init__.py
│   ├── logging.conf
│   ├── reference_config.json
│   ├── template_report.html
│   └── tool_config.json
└── utils
    ├── bam.py
    ├── config.py
    ├── fastq.py
    ├── __init__.py
    ├── logger.py
    ├── plot.py
    ├── report.py
    ├── sam.py
    ├── stat.py
    └── tag.py

Last news

• evatoolt 0.1.1 upload evatool to pypi
• evatoolt 0.1.2 add README.md
• evatoolt 0.1.8 fixed bug