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Version: 3.24 (unreleased)

Getting Started

Illumina Connected Annotations is written in C# using .NET Core (an amazing runtime environment that currently runs on Windows, Linux, Mac OS X, and in Docker images). Once .NET Core has been downloaded, all you need to do is grab the source, compile it, and grab the data files.

tip

Illumina Connected Annotations currently uses .NET6.0. Please make sure that you have the most current runtime from the .NET Core downloads page.

Getting Illumina Connected Annotations

Latest Release

Please visit Illumina Connected Annotations. to obtain the latest release.

mkdir -p IlluminaConnectedAnnotations/Data
cd IlluminaConnectedAnnotations
unzip IlluminaConnectedAnnotations-3.22.0-0-gc13dcb61-net6.0.zip

Quick Start

If you want to get started right away, we've created a script that unzips the Illumina Connected Annotations build, downloads the annotation data, and starts annotating a test file:

bash ./TestIlluminaConnectedAnnotations.sh IlluminaConnectedAnnotationsBuild.zip

We have verified that this script works on Windows (using Git Bash or WSL), Linux, and Mac OS X.

Docker

Obtain the docker image in a zip file (e.g. IlluminaConnectedAnnotations-3.22.0-0-gc13dcb61-net6.0-docker.tar.gz), and load it as follows

docker load < IlluminaConnectedAnnotations-3.22.0-0-gc13dcb61-net6.0-docker.tar.gz

If you want to build your own docker image, it is really easy to do. You just need to have Illumina Connected Annotations zip file and then download the Dockerfile and this script.

Put both files (create_docker_image.sh and Dockerfile) inside the same folder.

In terminal, execute command below inside the folder where you put those scripts:

chmod +x create_docker_image.sh
./create_docker_image.sh [path to zip file] [image tag]

After you run the script, the docker image will be available in your local machine with image name illumina-connected-annotations:[image tag specified].

For Docker, we have special instructions for running the Downloader:

docker run --rm -it -v local/data/folder:/scratch illumina-connected-annotations:v3.22.0 Downloader --ga GRCh37 -o /scratch

Similarly, we have special instructions for running IlluminaConnectedAnnotations (Here's a toy VCF in case you need it):

docker run --rm -it -v local/data/folder:/scratch illumina-connected-annotations:v3.22.0 Annotator -c /scratch/Cache/ \
     -r /scratch/References/Homo_sapiens.GRCh37.Nirvana.dat \
     --sd /scratch/SupplementaryAnnotation/GRCh37 \
     -i /scratch/HiSeq.10000.vcf.gz -o /scratch/HiSeq
caution

Please note that since our data files are usually accessed through a Docker volume, there is a noticeable performance penalty when running Illumina Connected Annotations in Docker.

tip

For convenience, the user is encouraged to create aliases for the docker commands. For example:

alias IlluminaConnectedAnnotations="docker run --rm -it -v local/data/folder:/scratch illumina-connected-annotations:v3.22.0 IlluminaConnectedAnnotations"

Downloading the data files

To download the latest data sources (or update the ones that you already have), use the following command to automate the download from S3:

dotnet bin/Release/net6.0/Downloader.dll \
     --ga GRCh37 \
     -o Data
  • the --ga argument specifies the genome assembly which can be GRCh37, GRCh38, or both.
  • the -o argument specifies the output directory
Glitches in the Matrix

Every once in a while, the download process does not go smoothly. Perhaps the internet connection cut out or you ran out of disk space. The Downloader attempts to detect these situations by checking the file sizes at the very end. If you see that a file was marked truncated, try fixing the root cause and running the downloader again.

tip

From time to time, you can re-run the Downloader to get the latest annotation files. It will only download the files that changed.

Preserving old data file

By default, while rerunning, the Downloader will replace old files with the latest versions. For example, if at some point, your SupplementaryAnnotation folder contained ClinVar_20231101.nsa and the latest available version is ClinVar_20231203.nsa, next time the Downloader is run, ClinVar_20231101.nsa will be replaced with ClinVar_20231203.nsa.

Currently, there is no way to override this behavior. If you do not want to replace/update any particular file, we recommend saving those files to a different location, rerun the Downloader to update the other data files and then manually replace the files you did not want updated. Please make sure to remove the latest version of the files you did not want. Note that the Annotator will throw an error if multiple versions of the same data source is present in the SupplementaryAnnotation folder. In other words, the SupplementaryAnnotation folder cannot contain both ClinVar_20231101.nsa and ClinVar_20231203.nsa.

Here is an example of how to proceed if a user doesn't want the latest version of ClinVar.

ls Data/SupplementaryAnnotation/GRCh38
...
ClinGen_disease_validity_curations_20231011.nga
ClinVar_20230930.nsa
ClinVar_20230930.nsa.idx
...
mv Data/SupplementaryAnnotation/GRCh38/ClinVar* <tmp_dir>/GRCh38/

dotnet bin/Release/net6.0/Downloader.dll \
     --ga GRCh38 \
     -o Data

rm Data/SupplementaryAnnotation/GRCh38/ClinVar*
mv <tmp_dir>/GRCh38/ClinVar* Data/SupplementaryAnnotation/GRCh38/

Download a test VCF file

Here's a toy VCF file you can play around with:

curl -O https://illumina.github.io/IlluminaConnectedAnnotationsDocumentation/files/HiSeq.10000.vcf.gz

Running Illumina Connected Annotations

Once you have downloaded the data sets, use the following command to annotate your VCF:

dotnet Annotator.dll \
     -c Data/Cache \
     --sd Data/SupplementaryAnnotation/GRCh37 \
     -r Data/References/Homo_sapiens.GRCh37.Nirvana.dat \
     -i HiSeq.10000.vcf.gz \
     -o HiSeq.10000
  • the -c argument specifies the cache directory
  • the --sd argument specifies the supplementary annotation directory
  • the -r argument specifies the compressed reference path
  • the -i argument specifies the input VCF path
  • the -o argument specifies the output filename prefix

When running Illumina Connected Annotations, performance metrics are shown as it evaluates each chromosome in the input VCF file:

---------------------------------------------------------------------------
Illumina Connected Annotations                      (c) 2023 Illumina, Inc.
                                                                     3.22.0
---------------------------------------------------------------------------

Initialization                                         Time     Positions/s
---------------------------------------------------------------------------
Cache                                               00:00:00.0
SA Position Scan                                    00:00:00.0      153,634

Reference                                Preload    Annotation   Variants/s
---------------------------------------------------------------------------
chr1                                    00:00:00.2  00:00:00.8       11,873

Summary                                                Time         Percent
---------------------------------------------------------------------------
Initialization                                      00:00:00.0        1.5 %
Preload                                             00:00:00.2        4.9 %
Annotation                                          00:00:00.8       18.5 %

Time: 00:00:04.4

The output will be a JSON file called HiSeq.10000.json.gz. Here's the full JSON file.

The Illumina Connected Annotations command line

The full command line options can be viewed by using the -h option or no options

dotnet Annotator.dll
---------------------------------------------------------------------------
Illumina Connected Annotations                      (c) 2024 Illumina, Inc.
                                                                     3.24.0
---------------------------------------------------------------------------

USAGE: dotnet Nirvana.dll -i <vcf path> -c <cache dir> --sd <sa dir> -r <ref path> -o <base output filename>
Annotates a set of variants

OPTIONS:
      --cache, -c <directory>
                             input cache directory
      --in, -i <path>        input VCF path
      --tsv <path>           input VCF path
      --out, -o <file path>  output file path
      --ref, -r <path>       input compressed reference sequence path
      --sd <directory>       input supplementary annotation directory
      --sources, -s <VALUE>  annotation data sources to be used (comma
                               separated list of supported tags)
      --credentialsFile <VALUE>
                             File path to user credentials, default is set to ~
                               /.ilmnAnnotations/credentials.json
      --ignoreLicenseError   ignore error due to invalid license and skip
                               related data sources
      --force-mt             forces to annotate mitochondrial variants
      --legacy-vids          enables support for legacy VIDs
      --enable-dq            report DQ from VCF samples field
      --enable-bidirectional-fusions
                             enables support for bidirectional gene fusions
      --disable-junction-preservation
                             disable junction preserving functional annotation
      --str <VALUE>          user provided STR annotation TSV file
      --vcf-info <VALUE>     additional vcf info field keys (comma separated)
                               desired in the output
      --vcf-sample-info <VALUE>
                             additional vcf format field keys (comma separated)
                                desired in the output
      --sa-cutoff <VALUE>    Any SV larger than or equal to this value will
                               not have any supplementary annotations
      --output-format <VALUE>
                             output file format, available options: json, vcf.
      --help, -h             displays the help menu
      --version, -v          displays the version

##### Supported Annotation Sources #####
Basic Tier: DECIPHER, GME, GERP, DANN, REVEL, ClinGen, gnomAD, phyloP, TOPMed, DGV, 1000 Genomes, CliinVar, dbSNP, FusionCatcher, MITOMAP, MultiZ100Way

Professional Tier: PrimateAI(GRCh37), PrimateAI-3D(GRCh38), SpliceAI, COSMIC, OMIM.

##### Contact #####
Professional content licensing, feedback and technical support: annotation_support@illumina.com.

Specifying annotation sources

By default, Illumina Connected Annotations will use all available data sources. However, the user can customize the set of sources using the --sources|-s option. If an unknown source is specified, a warning message will be printed.

dotnet Annotator.dll \
     -c Data/Cache/GRCh37 \
     --sd Data/SupplementaryAnnotation/GRCh37 \
     -r Data/References/Homo_sapiens.GRCh37.Nirvana.dat \
     -i HiSeq.10000.vcf.gz \
     -o HiSeq.10000 \
     -s omim,gnomad,ense
 ---------------------------------------------------------------------------
 Illumina Connected Annotations                      (c) 2023 Illumina, Inc.
                                                                     3.22.0
 ---------------------------------------------------------------------------

 WARNING: Unknown tag in data-sources: ense.
 Available values are: aminoAcidConservation,primateAI,dbsnp,spliceAI,revel,cosmic,clinvar,gnomad,
 mitomap,oneKg,gmeVariome,topmed,clingen,decipher,gnomAD-preview,clingenDosageSensitivityMap,
 gerpScore,dannScore,omim,clingenGeneValidity,phylopScore,lowComplexityRegion,refMinor,
 heteroplasmy,Ensembl,RefSeq

 Initialization                                         Time     Positions/s
 ---------------------------------------------------------------------------
 SA Position Scan                                    00:00:00.3      307,966
 ....
 ..

The list of available values is compiled from the files provided (using -c and --sd options).