Metadata-Version: 2.1
Name: nanome-jax
Version: 1.3.6
Summary: NANOME (Nanopore methylation) pipeline developed by Li Lab at The Jackson Laboratory
Home-page: https://github.com/TheJacksonLaboratory/nanome
Author: Yang Liu
Author-email: yang.liu@jax.org
License: UNKNOWN
Project-URL: Bug Tracker, https://github.com/TheJacksonLaboratory/nanome/issues
Platform: UNKNOWN
Classifier: Programming Language :: Python :: 3
Classifier: Intended Audience :: Science/Research
Classifier: License :: OSI Approved :: MIT License
Classifier: Operating System :: OS Independent
Description-Content-Type: text/markdown
License-File: LICENSE

# DNA methylation-calling tools for Oxford Nanopore sequencing: a survey and human epigenome-wide evaluation
## --NANOME(Nanopore methylation) pipeline of DNA methylation calling tools for Oxford Nanopore sequencing 

## Methodology of NANOME pipeline

**Background:** Nanopore long-read sequencing technology greatly expands the capacity of long-range, single-molecule DNA-modification detection. A growing number of analytical tools have been developed to detect DNA methylation from nanopore sequencing reads. Here, we assess the performance of different methylation calling tools to provide a systematic evaluation to guide researchers performing human epigenome-wide studies.


![Figure1A](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/Fig1A.jpg)

**Fig. 1A. Survey of methylation calling tools .**  Timeline of publication and technological developments of Oxford Nanopore Technologies (ONT) methylation calling tools to detect DNA cytosine modifications. 


![Figure1B](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/Fig1B.jpg)
**Fig. 1B. Workflow for 5-methylcytosine (5mC) detection for nanopore sequencing.** 


**Results:** We compared several analytic tools for detecting DNA modifications from nanopore long-read sequencing data. We evaluated the CpG methylation-detection accuracy, CpG site coverage, and running time using nanopore sequencing data across different genomic contexts, using natural human DNA. Furthermore, we provide an online DNA methylation database (https://nanome.jax.org) with which to display the DNA methylation levels detected by nanopore sequencing and bisulfite sequencing data across different genomic contexts.


**Conclusions:** Our study is the first benchmark of state-of-the-art methods for detection of mammalian whole-genome DNA-modifications in nanopore sequencing. We provide a broad foundation for cross-platform standardization, and an evaluation of analytical tools designed for genome-scale modified-base detection using nanopore sequencing. 

## System Requirements
### CI/CD features of NANOME
We use CI Automation Tools to enable the automated testing on every commit and on PRs to make sure that updates are not introducing bugs. Please check testing results on [Github](https://github.com/TheJacksonLaboratory/nanome/actions).

### Hardware requirements

NANOME is based on [Nextflow](https://www.nextflow.io/) pipeline framework, and start with raw fast5 nanopore sequencing input data with a reference genome. The pipeline can be configured with different RAM, CPU/GPU resources schema to parallelly run methylation-calling tools. For optimal usage, we recommend running NANOME pipeline on HPC or cloud computing platform, e.g., google cloud platform (GCP):
* GPU or CPU with 2+ cores. 
* RAM: 7+ GB per cpu.
* Storage using HDD or SSD. Please ensure your storage before running the pipeline.


### Software requirements
NANOME pipeline uses Nextflow technology. Users only need to install Nextflow and one of below commonly used environment tool:
* conda
* docker
* singularity

We provide conda, docker and singularity environments that depend on below well-known open-source packages for methylation calling on nanopore sequencing data:

[nanopolish](https://github.com/jts/nanopolish) >=0.13.2  
[megalodon](https://github.com/nanoporetech/megalodon) >=2.2.9  
[deepsignal](https://github.com/bioinfomaticsCSU/deepsignal) >=0.1.8  
[ont-tombo](https://github.com/nanoporetech/tombo) >=1.5.1  
[deepmod](https://github.com/WGLab/DeepMod) >=0.1.3  
[METEORE](https://github.com/comprna/METEORE) >=1.0.0  
[ont-pyguppy-client-lib](https://github.com/nanoporetech/pyguppyclient) >=4.2.2  
[fast5mod](https://github.com/nanoporetech/fast5mod) >=1.0.5

Guppy software >= 4.2.2 from [ONT (Oxford Nanopore Technologies) website](https://nanoporetech.com)


## Installation
Users only need to install **Nextflow**, see [installation document](https://www.nextflow.io/docs/latest/getstarted.html#installation). NANOME execution environment will be automatically configured with the support of conda, docker or singularity containers.

NANOME pipeline support running with various ways in different platforms:
* Conda
    1. Create conda enviroment: `conda env create -f environment.yml`
* Docker
    1. Docker container name from [Docker Hub](https://hub.docker.com/repository/docker/liuyangzzu/nanome): `liuyangzzu/nanome:latest`, you can also build docker image by `docker build -t liuyangzzu/nanome:latest .`
* Singularity
    1. Pull image from [Docker Hub](https://hub.docker.com/repository/docker/liuyangzzu/nanome): `singularity pull docker://liuyangzzu/nanome:latest`
* HPC clusters with **SLURM** support
* Google Cloud platform with **google-lifesciences** support

## Simple usage
NANOME pipeline can be directly executed without any other additional installation steps:
```angular2html
# Run NANOME using docker
nextflow run TheJacksonLaboratory/nanome\
    -profile ci,docker

# Run NANOME using singularity
nextflow run TheJacksonLaboratory/nanome\
    -profile ci,singularity
```

Please refer to [Usage](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/Usage.md) for how to use NANOME pipeline. For running on CloudOS platform (e.g., google cloud), please check [Usage on CloudOS](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/Usage.md#4-running-pipeline-on-cloud-computing-platform). 

If you prefer using our evaluation packages, please check [evaluation script usage](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/Eval.md) for more detail.

## Pipeline reports for NANOME
### Benchmarking reports on our HPC using [Nextflow](https://www.nextflow.io/)

We constructed a set of benchmarking datasets that contain reads from 800 to about 7,200 reads for NA19240, and monitored job running timeline and resource usage on our HPC, reports generated by **Nextflow** workflows are: [Trace file](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/resources/trace_benchmark.txt.tsv), [Report](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/resources/report_benchmark.pdf)  and [Timeline](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/resources/timeline_benchmark.pdf). 

Our HPC hardware specifications are as follows:
* CPU: Intel(R) Xeon(R) Gold 6136 CPU @ 3.00GHz
* GPU: Tesla V100-SXM2-32GB 
* RAM: 300 GB
* Slurm manager version: 19.05.5

Timeline figure for benchmarking experiments are below:
![Bench-timeline](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/resources/timeline_benchmark.jpg)


### Pipeline DAG
![NanomeDag](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/nanome_dag.png)

### NANOME report
Please check [NANOME report](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/NANOME_report_html.pdf) for the sample report by NANOME pipeline.

![NanomeReportHtml](https://github.com/TheJacksonLaboratory/nanome/blob/master/docs/nanome_report_html.png)



## Revision History

For release history, please visit [here](https://github.com/TheJacksonLaboratory/nanome/releases). For details, please go [here](https://github.com/TheJacksonLaboratory/nanome/blob/master/README.md).

## Contact

If you have any questions/issues/bugs, please post them on [GitHub](https://github.com/TheJacksonLaboratory/nanome/issues). We will continuously update the Github to support famous methylation-calling tools for Oxford Nanopore sequencing.

## Reference
Detailed results can be found in our publication. Please cite our article below if you are interested in our GitHub repository:

**DNA methylation-calling tools for Oxford Nanopore sequencing: a survey and human epigenome-wide evaluation.** Genome Biology 22, 295 (2021). https://genomebiology.biomedcentral.com/articles/10.1186/s13059-021-02510-z 


