The Top navigation panel of the platform serves as a guide to the platform. It contains:

• Dashboard, Cases and Add new case buttons lead to the corresponding pages

• Case search bar

• Settings dropdown menu activated by clicking on the user name or profile picture

• Help dropdown menu activated by clicking on the question mark icon

• Emedgene Applications menu activated by clicking on the app launcher icon on the left

As of version 2.26:

Emedgene is an AI-based platform, incorporating machine learning throughout the analysis and interpretation workflow in order to deliver the fastest time from genomic data to decisions. We apply machine learning models that retrieve evidence-backed answers and provide exceptional decision support.

• The platform is not a black box, and overlays a layer of explainable AI (XAI), presenting supporting evidence from the literature and databases which significantly reduces the time to interpret a case.

• The algorithms use a proprietary Emedgene knowledge graph which incorporates information extracted from literature with Natural Language Processing, as well as from public databases and is updated on a monthly basis.

• Dozens of additional algorithms are incorporated throughout the workflow.

Overall, the system combines AI in a highly optimized and customizable workbench, in order to automate the most time-intensive aspects of genomic analysis and research.

The Emedgene platform is divided into two applications:

• Analyze - genomic analysis workbench,

• Curate - the knowledge management system.

To switch from Analyze to Curate:

Go to the nine-dot app launcher icon located on the left of the Top navigation panel and select Curate from the dropdown menu.

To switch from Curate to Analyze:

Go to the nine-dot app launcher icon located on the Curate navigation panel and select Analyze from the dropdown menu.

The Emedgene platform utilizes the Okta Identity Management solution to control user access. This improves user management, enhances access and authentication security, and allows organizations to implement single sign-on for their users.

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To streamline case review, the AI Shortlist pre-selects the list of variants likely to be causative for each case:

Most Likely Candidates

Variants that are most promising for solving the case. This list is limited to 10 top-scored variants but may include more if more than one variant is tagged per gene (suggesting compound heterozygosity). We can change the Most Likely Candidates number limit upon request.

Candidates

Several dozen highly scored variants worth considering.

As of version 30.0 and onwards, the ranking of variants by AI Shortlist considers both SNVs and CNVs, including SNV + CNV compound heterozygotes. Starting from version 32.0, AI Shortlist additionally considers SVs, mtDNA variants and STRs.

The AI Shortlist rates variants based on predicted variant effects, alternative allele frequency, familial segregation pattern, phenotypic match, in silico predictions, and other relevant information from scientific papers and databases.

During the case review, you can untag variants selected by the AI Shortlist or manually tag ones not selected by the AI Shortlist.

Welcome to Emedgene, where we unlock genomic insights for hereditary disease and streamline your tertiary analysis workflows.

So you've signed in and can't wait to get started? Here we will guide you through the platform architecture, case creation, and results review. You can dive a bit deeper by following the links and exploring manuals for the platform's applications:

• Analyze - Genomic analysis workbench, where you can accession, interpret, curate and report on your cases, while also efficiently managing the lab workflow.

• Curate - A repository for all of your organizational curated knowledge.

Look around

The platform is operated from the Top navigation panel.

By clicking on the corresponding buttons, you can enter:

• Cases page

• Add new case page

• Emedgene Applications menu

• Help dropdown menu

• Settings dropdown menu

Create a case

To enter the Add new case flow, click on the namesake button on the Top navigation panel. Here:

• Select file type,

• Upload files,

• Create a family tree,

• Annotate each sample with clinical information,

• Specify analysis details, and

• Launch the analysis!

You'll be notified when results are ready.

Examine the analysis results

Select a case to review on the Cases page. You'll be directed to the Individual case page that:

• Showcases an AI-curated shortlist of variants suggested to be checked first, namely Most Likely Candidates and Candidates,

• Provides numerous customizable filters to help you explore the total list of genetic variants by yourself, and

• Documents all the case-related information like Case status, sample quality metrics, and versions of all the resources used during case analysis.

On the Variant page, each variant can be thoroughly investigated and accordingly tagged.

When you're ready to finalize the case, indicate the end result of the analysis and variants to be reported in the Case interpretation widget.

Click on the question mark icon of the Top navigation panel to open the Help dropdown menu.

From there, you can access:

• Help Center. Feeling curious? Dive right in.

• What's New. Check out the release notes to be on top of our latest and greatest features!

Whenever an organization is created, we automatically allocate bucket folders in AWS S3 cloud storage to it:

• Path for upload

<Region_Cloud>-emg-auto-samples/<org_name>/upload/

Folder intended to store input case files.

Authorized user has view and upload privileges.

• Path for download (32.0+)

<Region_Cloud>-emg-downloads/<org_name>/ 

This folder contains a partially annotated (excluding results of proprietary algorithms) VCF file per case.

Authorized user has view and download privileges.

• Path for DRAGEN output (32.0+)

<Region_Cloud>-emg-auto-results/<org_name>/ 

This folder contains DRAGEN output files.

Authorized user has view and download privileges.

To get access to your upload, download and DRAGEN output folders, you need to get a key pair consisting of an access key ID and a secret access key. Creating, deactivating, activating and deleting credentials is available for users with Manager and Manage S3 Credentials roles.

You can create and use up to two dynamic access keys at the same time.

When you require technical support, you have the option to generate a new key pair specifically for the troubleshooting process. After the issue has been resolved, you can delete the credentials to ensure security of your system.

The newly generated credentials will only be saved in AWS Identity and Access Management (IAM) and not in our database.

How to create a key pair

1. In Settings > Management > S3 Credentials, click on Create Access Key.

2. You can retrieve the secret access key only when you initially create the key pair. If you lose it, you have to create a new key pair. To immediately copy the secret access key to a secure location, use the Copy to clipboard button.

How to deactivate a key pair

In Settings > Management > S3 Credentials, click on Deactivate in the corresponding key pair card.

How to activate an inactive key pair

In Settings > Management > S3 Credentials, click on Activate in the corresponding key pair card.

How to delete a key pair

In Settings > Management > S3 Credentials, click on Delete in the corresponding key pair card. Only inactive key pairs can be deleted.

Version 33.0+:

With version 33.0 and later, you have the flexibility to manage Case labels at any time: create, add, or remove them directly in the Cases table.

Versions older than 33.0:

The organization's custom Case labels must be assigned while creating a case, in the Case info screen of the Add new case flow.

The desired Case labels should be created prior to case creation by the organization's manager in the Organization settings.

Once a case is created, Case labels cannot be removed or added.

The Analysis tools tab contains multiple tools that facilitate a case review:

• Filters panel - includes numerous adjustable Filters and Presets (on the left)

• Variant table - a user-customizable display of all the analysis results (core section)

• Variant page - when selecting a variant, the variant page opens, revealing a detailed annotation and other assessment tools for the particular variant.

The Zygosity Filters enable manual selection of zygosity status (Het, Hom, Ref, No Cov) for each sequenced sample in the pedigree.

The Dashboard provides a glance at key performance indicators for an organization and depicts an overview of the user activity on the Emedgene platform.

This page is divided into two panels and contains the following subsections:

Lefthand panel

• Emedgene Knowledge Base panel reveals the total amount of known polymorphisms, pathogenic variants, and gene-disease connections considered during analysis. It also indicates the number of scholarly publications processed by Emedgene's novel textual recognition platform, backed by natural-language processing and artificial intelligence.

• Diagnostic Yield panel presents the proportion of "solved" cases out of the total number of the organization's cases of the same type.

• Status Diagram panel displays the total number of the organization's submitted cases as well as the numbers of cases under each status.

• Stale Cases panel highlights the cases that are stuck at one of the intermediate stages of the analysis, and are not finalized.

Righthand panel

The Cases tab provides a bird's-eye view of all the previously submitted genomic sequencing cases.

The Cases tab includes:

Cases table lists all the genomic sequencing cases submitted by your organization. It is the primary component on the left-hand side of the Cases tab.

Brief column description:

Case ID

A unique ID assigned to a case by Emedgene.

Proband ID

Proband's sample ID submitted by creating the case in the platform.

Status

Creation Date

Automatically saved.

Due Date

Can be set, changed or removed on the spot. To set a date, click on the calendar icon and select a date of interest. To change it, click on the previously set due date and select a new one. To remove the due date, click on the cross button next to it.

Phenotypes

Proband phenotypes as submitted by the user.

Participants

Users involved in the case submission, analysis, finalizing, or those who subscribed to receive updates on the case. To receive email notifications on your colleagues' activities in the particular case, click on Subscribe icon. To unsubscribe, hover over your initials and click on the cross button.

Type

Case Type (Array, Custom Panel, Exome, Whole Genome).

Label

Dedicated user groups

Family tree legend:

1. Icon fill color in other pedigree members indicates the presence or absence of the proband's phenotypes in a present sample (regardless of the potential presence of additional unrelated phenotypes):

2. Icon color intensity denotes whether sample files have been uploaded for the particular individual:

3. Icon line type indicates whether the sample is considered or excluded during analysis (relevant to samples with uploaded files only):

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The Individual case page includes:

Highlights a shortlist of variants, suggested to be reviewed first - Most Likely Candidates and Candidates.

Illustrates quality metrics for the sequenced samples.

Provides numerous customizable filters to help you explore the total list of genetic variants in compliance with your organization's standard case review process. You can export shortlisted variants in .xlsx format.

Documents versions of all the resources used during case analysis.

The Variant Type Filters allow filtering variant list by variant type:

• Sequence variants: SNV, Indel (Note: Indel filter requires pipeline version 5.22.8 or higher);

• CNV: DEL, DUP;

• mtDNA SNV/Indel.

Examples of CNV overlap percentage calculation (see legend below):

• Orange represents the current CNV;

• Violet blue represents a previously reported CNV;

• Darker shade indicates the region of overlap between the current and previously reported CNV.

Options available through the Top bar:

• Reanalyze the case

Case details panel is divided into three tabs:

• Case Info

• Family Tree

• Activity

How to see case details

Information on the currently selected case is displayed in the window that pops up when you click on the corresponding row of the Cases table. To close the window, click on the cross icon.

Information presented on the Case details panel:

Case Info

1. Technical details:

1. Case ID

2. Case Type: Custom Panel, Exome, Whole Genome

3. Sample Type: FASTQ, Project VCF, VCF, BAM

4. Gene List - all genes or a particular gene list used to filter the analysis results

5. Human Reference - the genome reference used during case analysis

2. Operational details:

1. Ordered by - user who created the case by default, and creation date

2. Signed by - user who finalizes the case

3. Related cases - lists the Case IDs for all the cases that share one or more samples with the one currently selected

4. Due Date - a deadline for finalizing the case. You can enter or edit the Due Date by clicking on the calendar icon under the Due Date section.

5. Participants - names of the users involved in the case submission, analysis, finalizing, or those who subscribed to receive updates on the case. To receive email notifications on your colleagues' activities in the particular case, click on the Subscribe icon.

3. Clinical details:

1. Patient Information: Sex (33.0+) / Gender (32.0 and older), Age

2. Clinical Information:

   1. Proband Phenotypes - HPO terms used to describe clinical findings in the proband

   2. Suspected Disease: Suspected disease (if provided), Penetrance (%) and Severity (mild, moderate, severe, or profound)

   3. Maternal and Paternal ethnicity

   4. Parental Consanguinity

   5. Report secondary findings (Yes, No or N/A)

3. Clinical Note: any notes on proband's phenotypes, family history, or other critical points of the case.

Family Tree

Here you can find:

1. Graphic representation of the pedigree. More information about the symbols can be found here.

2. Sample information for each family member:

   1. Phenotypes: proband phenotypes and phenotypes reported for other family members (related and unrelated)

   2. Medical Condition (Healthy or Affected)

   3. Sex

   4. Age

   5. Maternal and Paternal ethnicity

   6. BAM file location

Activity

This tab logs actions related to the selected case such as Case status changes, variant tagging, ACMG pathogenicity, changes to an evidence graph, evidence notes, transcript changes for a specific variant, and comments added by users. Each log includes the date and time that each action was performed. We keep all logs for at least six years for full traceability.

You can choose All activities, Comments, or Case-related activities from a dropdown menu. To add a comment to the case, write it in the Write a new comment text field and click Add.

The Candidates tab presents:

Most Likely Candidates and Candidates

A shortlist of the most promising variants with the highest scores from the AI Shortlist:

• before 30.0: SNVs;

• 30.0+: SNVs and CNVs;

• 32.0+: SNVs, CNVs, SVs, mtDNA variants and STRs.

These variants are initially selected by the AI Shortlist, but you may untag variants or tag them manually during the case review.

Incidental

Pathogenic or likely pathogenic variants in the medically actionable genes defined by the ACMG. These variants are automatically tagged only if you've selected the Secondary findings checkbox while creating a case.

Carrier

Variants identified by the Carrier analysis pipeline. Carrier variants are automatically tagged only if you've selected the Carrier Analysis checkbox while creating a case. Analysis requirements and a list of targeted regions are specified by the organization's manager. This Carrier analysis flow is implemented by request.

In Report and other custom variant tags

Variants that were manually selected to be reported.

Reviewing the Candidates tab

To select variants with a particular tag, use the Filter candidates dropdown menu in the top right corner. You can choose between Most Likely, Candidate, Incidental, Carrier, Not Reviewed, or any custom tags used in your organization.

For each variant on the Candidates tab, you can explore the suggested diagnosis, gene symbol, main variant details, and variant tag.

When a variant is found in a gene with no known association with a disease, the possible diagnosis cannot be indicated. Such variants are displayed under the Gene of Unknown Significance title.

All the relevant Most Likelies and Candidates fitting a сompound heterozygous mode of inheritance are presented together. This refers to both confirmed and assumed compound heterozygosity (cases with at least one parent and singleton cases, respectively).

If you want to inspect the complete variant information, click on the variant bar to continue to the Evidence page. You can visualize evidence in text or graphical format (Click on the interactive text in the top left corner: Show evidence as text or Show evidence graph to toggle between the two).

You can search for variants in the Variant search box by:

Single phenotype

"phenotype1" (e.g., "Mandibular prognathia")

Disease

"disease1" (e.g., Kabuki syndrome 1)

Disease inheritance mode

"inheritance mode1" (e.g., Autosomal dominant);

Genomic position

"coordinate1" (e.g., chr11:2686616)

Specific SNV/indel variant (32.0+)

"variant" (e.g., chr1:27089776G>T)

Genomic range

"range1" (e.g., chr11:2686616-2886620)

CNV length

"cnv_size:size1" (e.g., cnv_size:100000-10000000)

Gene symbol

"gene1" (e.g., BRCA1)

Multiple gene symbols added in batch

"gene1, gene2, gene3" (e.g., BRCA1, BRCA2, UBE3A)

Most of the above-listed options may be combined.

If you would like to update or make corrections your case details, phenotypes or gene list, you have the option to edit the case and save or reanalyze the data.

How to edit case data:

1. Open a case,

2. Press Edit case info button in the top right corner,

3. You'll access the Add new case flow. You can edit any information on the Family tree screen, as well as Select genes list, Select preset, and Additional case info sections of the Case info screen.

1. After you've finished editing the case and pressed Next in the Case info screen, a window will pop up:

While navigating between variants in the emedgene platform, there is an option to change the genomic position in the full-featured IGV desktop application according to the currently selected variant. How cool is that?

In order to enable or disable control of IGV from a web browser, please open your desktop IGV application and follow the instructions:

1. Go to the View menu and select Preferences.

1. Go to the Advanced tab.

1. Select or unselect the Enable port option to enable or disable the feature, respectively.

1. Save the changes.

That's it!

The Desktop apps panel allows you to activate or inactivate connections with your IGV and Alamut desktop applications. This means there won't be any IGV and Alamut windows popping up unless you want them to!

Once you've configured your preferences for the desktop applications connections, they will be saved for your user account and applied to all cases.

The Desktop apps panel appears as a tab in the Variant page sidebar.

Individual case page > Analysis tools tab > Variant table > Variant page

The Variant page showcasing the comprehensive variant information is accessible from the Variant table by selecting the corresponding variant row with a click. Once you're on the Variant page, you can move between variants using left and right arrow keys of your keyboard.

The Variant page is comprised of:

1. Top bar. Displays Case ID, gene symbol, genomic DNA-level description of the variant, variant tags, and a link to your Curate database. If the variant is already in your Curate database, you will see an Open Curate button. Otherwise, you will see an Export to Curate button.

2. Navigation panel (left). Divided into five tabs, leading to the corresponding page sections.

3. Page body:

   1. Summary section. Highlights core variant-related information from other sections

   2. Clinical Significance section. Reports essential variant- and gene-level information and indicates gene-related diseases.

   3. Quality section. Outlines the major variant quality parameters in each sample and demonstrates the family tree with the zygosity for each sequenced sample.

   4. Visualization section. Features the IGV-based BAM file viewer.

   5. Population Statistics section. Addresses alternative allele frequency, alternative allele count, and the number of homozygotes in public and internal databases.

   6. Related Cases section. Displays statistics regarding the pathogenicity and tags assigned to the variant under review, incorporating data from previous cases within both your organization and networks.

   7. Evidence section. Highlights user-selected variant pathogenicity, ACMG class (for a sequence or a genomic variant), and interpretation notes.

4. Variant activity panel (right). Records variant-level user activities, such as tagging a variant, adding comments or evidence notes, or editing the evidence graph. Variant activity panel pops up upon clicking the Activities button.

The Quality section:

1. Demonstrates the family tree with zygosity status and the overall variant quality indicated for each sample. Zygosity (HET, HOM, HEMI, or REF) is marked inside the pedigree symbol, and variant quality grade (H, M, or L) is denoted on the side.

1. Outlines the major variant quality parameters underlying the general grade in each sequenced individual:

• SNV/Indel variants:

  • Base Quality,

  • Depth,

  • Mapping Quality,
• CNVs:

  • Copy Number,

  • CNV Quality,

  • Size,
• STRs:

  • Depth;

  • Repeat Number,

1. Illustrates the allele fraction per sample in a pie chart. Not relevant for CNVs.

To change the sample in review for the particular variant, click on the corresponding icon in the family tree.

The Versions tab reports versions of all the tools and resources used during the case analysis in the following categories:

• Annotation

• Emedgene Resources

• Knowledgebase Sources

• Organization Local Databases

• Population Databases

• Variant Databases

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Add new case page > Family tree screen > Create family tree panel

Build a pedigree via the visual tool.

It is ideal that a proband selected for case analysis is affected and has disease phenotype(s).

You can add a Father, a Mother, a Sibling, or a Child to any family member, starting with the Proband. To do this, choose their icon, then click on the Add family member button in the bottom right corner of the pedigree builder to select a family member.

More information about the pedigree symbols can be found here.

To delete a family member, choose their icon, then click on the Delete Subject button in the top right corner of the Add patient information panel.

On the Evidence page, each proband's phenotype is marked according to the degree of similarity to the phenotypes observed in the genetic condition presumably associated with a variant under review.

Phenotypic match strength levels:

Exact match

Same term as in the clinical synopsis of the suspected disease

Match by ascendance

Phenotypes share a parent term in the HPO hierarchy

Indirect match

Phenotypes are closely related in the HPO hierarchy, but term relatedness is lower than in Match by ascendance

Unmatch

The phenotype is not reported for the suspected disease.

Each phenotypic match strength level is denoted by a particular icon next to the HPO term in the Case info tab of the Case details panel:

The Population Statistics section addresses detailed population allele data across various ethnicities in public and internal databases.

• Public databases (SNVs): 1000 Genomes, ESP 6500, ExAC, and gnomAD

• Public databases (CNVs): 1000 Genomes, gnomAD SV, Decipher, DGV

• Internal databases: EmedgeneDB and organization's NoiseDB or other custom databases

By clicking on a particular row of the table, it will provide additional details including alternative allele frequency, alternative allele count, and homozygotes count reported for the selected population by different sources.

Population Statistics for mtDNA variants

The Population Statistics section displays population allele data across various ethnicities in:

• Public databases: gnomAD and MITOMAP

• Internal databases: EmedgeneDB and organization's NoiseDB or other custom databases

By clicking on a particular row of the table, it will provide additional details including the highest homoplasmy frequency, heteroplasmy count, homoplasmy count, and the total number of samples.

Visualization is immediately accessible for cases that have been analyzed from FASTQ or BAM files. If the analysis was performed from VCF files, mapping visualization can be enabled using local BAM files.

In a VCF case, read alignment view can be enabled in two ways:

a. Creating a case via API with location of alignment files* defined in JSON;

b. Loading local alignment files* to your case in Emedgene.

Click on the button in the top right corner of the section and select relevant BAM and BAI files stored on your PC. After that, you will be able to select the samples to be shown in the viewer by selecting the corresponding sample names on top of the section.

• BAM and BAI (or CRAM and CRAI)

Add new case page > Family tree screen > Create family tree panel > Show Secondary Findings

While creating a case, you can choose if you want Secondary (Incidental) findings in the Proband to appear in the results.

These are known or expected pathogenic variants in the genes that are unrelated to the primary purpose of the testing. The secondary finding genes list includes:

• 81 genes (Miller et al. 2023) on versions 33.0+;

• 78 genes (Miller et al. 2022) on versions <33.0.

To directly import files from your storage, link storage to your organization in Emedgene.

How to create a link between storage and organization:

1. Select the Management tab. Under Storage is a list of currently linked storages. To add a new one press on Add Storage button.

1. Choose a Storage Type from:

   1. Azure Data Lake;

   2. Azure Blob;

   3. AWS S3;

   4. File Transport Protocol (FTP);

   5. Secure File Transport Protocol (SFTP);

   6. Illumina Basespace (BSSH);

   7. Illumina Connected Analytics (ICA).

1. Fill in the required credentials.

2. Click on Add storage:

1. Check the connection to confirm that the storage is successfully linked.

To do this, find the storage in the Storage List and check the cloud icon next to its name: 1. If it's green, the connection is set correctly; 2. If it's red and strikethrough, something went wrong. Hover over the icon to see details.

How to edit storage information:

In the Storage List press Manage on the right to the storage details.

How to remove a link to storage:

In the Storage List press Delete on the right to the storage details.

Prerequisites

• Emedgene platform version >= 32

Batch upload via CLI (Command Line Interface)

1. Download the batch case create script. Replace my-domain with your Emedgene domain. Illumina cloud: my-domain.emg.illumina.com Legacy Emedgene cloud: my-domain.emedgene.com

1. Download the CSV template file.

2. Execute the batch cases creator as java script using the command below. Replace my-domain with your Emedgene domain and my-email with your user email. A prompt for your Emedgene password will appear, enter the password and press Enter.

1. In case of validation errors in the input CSV, an output CSV called batchCases_results.csv will be created in the same location with detailed error results.

2. -l will create a log file in the same location.

More information can be found by running

The Genome View Tab, available from V37.0 and up, is a powerful feature designed to give users a clear, visual overview of the genome and chromosomes in their cases. This feature is especially useful for analyzing large Copy Number Variation (CNV) events and regions of homozygosity/loss of heterozygosity (ROH/LOH) across the genome, providing intuitive filtering and interactive insights for researchers and clinicians.

What is the Genome View Tab?

The Genome View tab is a dedicated section within the Case Page for Whole Genome Sequencing (WGS), Whole Exome Sequencing (WES), and Array data. This tab offers a graphical visualization of genomic data, focusing on CNV and LOH/ROH analysis for proband cases. Users can access this tab directly from the Case Page.

Key Features

Chromosome Ideogram

The chromosome ideogram offers a visual representation of all 23 human chromosomes, with CNV and LOH/ROH events highlighted for intuitive analysis. Here's what you can expect:

Variant types:

• Deletions (DEL): Marked in red, displayed to the left of the chromosome.

• Duplications (DUP): Marked in blue, displayed to the right of the chromosome.

• LOH/ROH (Regions of Homozygosity/Loss of Heterozygosity): Marked in gold, displayed over the chromosome.

Note: Variants with no coverage or reference (Ref) are excluded.

Filtering Options:

• Users can filter segments by size, using a range selector with the following options: 50 bp,1 KB, 10 KB, 50 KB, 100 KB, 1 MB, 10 MB, Max (no filter).

• The default filter is set from 50 KB to Max.

• Limitation: Only the 500 largest variants are displayed in this tab in v37.0.

Hover and Click Interactions:

• Hovering over a variant displays

  • Chromosome number, start and end positions, and size (e.g., chr1:100000-200000 (100 KB))

  • Cytoband range (e.g., p12.3 - q11.2) based on ISCN nomenclature

  • Variant type (DEL/DUP/LOH/ROH)

  • Number of genes affected

• Clicking on a chromosome refines the genome view below to that chromosome.

• Clicking on a variant opens a detailed Variant Page where many actions and further review can be made.

• Legend: A clear legend explains color coding and icons for DEL, DUP, and LOH/ROH variants for quick reference.

Genome Viewer

The genome viewer provides a deeper dive into the genomic data through three interactive tracks:

• Log R / TNS Track:

  • Displays copy number intensity data using values from the TNS BigWig file or LogR bedgraph.

  • Y-axis ranges from -3 to 3, with increments of 0.4.

  • X-axis displays the genome (whole genome view) or chromosome segments (whole chromosome view).

• BAF Track:

  • Displays B Allele Frequency (BAF) data using values from the BAF BigWig file or BAFbedgraph.

  • Y-axis ranges from 0 to 1, with increments of 0.1.

  • X-axis aligns with the Log R track.

• ROH Track:

  • Indicates regions of homozygosity for further analysis.

Zoom and navigation

• Default View: Displays the entire genome.

• Zoom-In Options: Users can zoom in to view individual chromosomes by clicking on them.

• Interactive Navigation: Clickable chromosomes on the ideogram allow seamless switching between views.

The Variant Effect Filters allow filtering variants by consequence, ACMG pathogenicity classes, and whether/how the variant has been classified internally or in clinical variation databases. The filters can operate in a Simple or Advanced mode.

Modes

Simple

Filter variant list by:

• Severity of variant effect (High, Moderate, Low, Modifier);

• CNV Severity is set according to the image below;

• Known Variant (Known Variants, Known Pathogenic Variants) - variant status in clinical variation databases (ClinVar) and previous classifications by your organization or network.

Advanced

Further restrict analysis results by:

• Specific Main effect of the variant on protein structure and function;

• ACMG Classification (Pathogenic, Likely Pathogenic, Uncertain Significance, Likely Benign, Benign)- ACMG pathogenicity class assigned manually or automatically. Note: applicable only for Candidate and Most Likely variants;

• ClinVar Known Variants (Pathogenic, Likely Pathogenic, VUS, Likely Benign, Benign, Other) - variant status in ClinVar;

• Custom database Known Variants (Pathogenic, Likely Pathogenic, VUS, Likely Benign, Benign, Other) - variant status in your own curated variant database, including CNVs detected by means of NGS and/or chromosomal microarray;

• Manually Classified Variants - select this option to restrict results to variants from previous cases with user-assigned Pathogenicity.

Default values

When Filters are Reset to Default, the Variant Filters are set to:

• Severity: High, Moderate, Low;

• Known Variant - no filtering.

Where can I review and edit the ACMG Classification of a SNV?

How do I review and edit the ACMG Classification of a SNV?

Each ACMG tag is represented by an interactive button including checkbox (1), name (2) and evidence strength indicator (3).

Pathogenic criteria are represented by red boxes, while benign criteria boxes are colored green. Each ACMG criterion has three possible states:

• Neutral (1) - represented by an empty checkbox. Criterion requires further investigation.

• Negative (2) - represented by a cross. Criterion is not applicable.

• Positive (3) - represented by a tick and dark color. Criterion is applicable.

Each ACMG tag can be manually checked, unchecked, or set to an undefined state by clicking the interactive button's checkbox element.

To examine in detail or modify the underlying evidence for the particular ACMG tag, select it by clicking on the tag name. The button becomes flood-filled (b), as opposed to it's original, non-selected, state (a).

Upon selection, a description of the criterion and its underlying evidence emerges below. Yes and No radio buttons accompany each piece of evidence. The tag can be assigned if Yes has been selected for all the underlying conditions.

You may modify evidence strength in the Strength dropdown (Stand Alone, Very Strong, Strong, Moderate, Supporting), which will impact both the pathogenicity class and score calculations.

On versions 32.0+, you have the capability to add a note alongside a tag.

After you've modified ACMG classification, you can either save manual changes by pressing the Save button or reset via Revert manual changes. Keep in mind that after saving your edits, Revert manual changes will become unavailable.

ACMG classification of mtDNA variants

The ACMG SNV Classification wizard is available for ACMG classification of tagged mtDNA variants. To classify an mtDNA variant, please manually assign the relevant criteria; the resulting ACMG classification will be calculated automatically.

For versions prior to 34.0, technical support is required to implement a customized filter Preset. But with version 34.0 and later, users can easily do it on their own.

Create filter Presets from active filters (34.0+)

To save Presets from active filters:

2. Click on the three-dot icon;

3. Select Save as preset;

4. Enter a name for the Preset (Note: Avoid using non-Latin symbols that don't follow the ISO-5589-1 standard);

5. Click Save.

Where can I manage Presets? (34.0+)

1. In Presets, scroll down and click Add beside Gene Lists.

1. Select the gene lists you want to utilize as filter presets by marking the relevant checkboxes. Starting from version 30.0+, a search bar simplifies list navigation by allowing you to search by list name.

1. Click Save.

Presets originating from gene lists will appear in Presets under Gene Lists.

Review logic behind the Preset (32.0+)

On 32.0+, for a quick refresher, you may review the logic behind each Preset directly within your analysis flow. To do so, click on an downward arrow icon left to the Preset's name.

You can select the sample's file type from the given options:

1. FASTQ: .fastq.gz, .fq.gz, .bam, .cram.

2. Project VCF: .pvcf, .vcf, .vcf.gz, .pvcf.gz

3. VCF: .vcf, .vcf.gz, .targeted.json, *.gt_sample_summary.json

You can choose one of the following options:

1. Existing sample - pick one of the samples already loaded on the platform

2. Upload New Sample - upload files from your PC and enter sample name

3. Choose from storage - choose files from your cloud storage and enter sample name

4. No sample - postpone uploading files but proceed with case creation or skip uploading files for family members other than Proband

Any variant can be tagged by the AI Shortlist or a user as:

• Most Likely Candidate - most promising for solving the case;

• Candidate - worth considering;

• Carrier - identified by the Carrier analysis pipeline;

• In Report - manually selected to be reported;

• Not relevant - automatically tagged variant that has been disregarded after manual review;

• Any custom tag used in your organization (e.g., Submitted for Sanger confirmation).

Variant tagging widget how-to

Add or change a tag

Click on the dropdown icon in the Variant tag field and select a suitable tag. The Variant tag field showcases the most recently assigned tag.

Review the tagging history

• 25.0.0+: click on the Variant tag field and scroll to the Assigned tags section.

• 23.0.0 and older: click on the View all tags.

Remove an automatic tag

Click on the dropdown array in the Variant tag field and select Not relevant.

Clear your manual tag:

Click on the dropdown array in the Variant tag field and select Clear.

Variants can be filtered by:

In the Filters tab of the Filters/Presets panel, the vertical ellipsis button offers options:

• Clear - deselect all filters and reveal all the variants;

• Reset to Default - set default filters values: moderate/high quality (Quality Filter), low/moderate/high severity (Variant Filters), and Het/Hom zygosity in the proband (Zygosity Filters);

The Polymorphism Filters enable filtering variants by alternative allele frequencies and genotype counts in public and internal databases. The filters can operate in a Simple or Advanced mode.

Modes

Simple mode

Switch on or off Display Polymorphism option:

• When switched on, no restrictions are being applied.

• When switched off, variants with allele frequency >0.05 in public databases or >0.25 in the EmedgeneDB are filtered out.

Advanced mode

Filter variant list by limiting the maximum Allele Frequency, Hom/Hemi and Het counts in one of the public database options (GnomAD, ExAC, 1000 Genomes, or GME) or, by default, in all (All Databases). The default values for Hom/Hemi and Het can be exceeded using the text box.

In addition, limit results by the maximum Allele Frequency in internal databases:

• Emedgene Database - a static sample set of 806 healthy individuals. Aimed at getting rid of the artifacts generated by our FASTQ processing pipeline

• Organization Databases, e.g., NoiseDB - a blacklist of variants (implemented by request).

Default values

You can expand to a broader genetic testing option if the results of more targeted testing are inconclusive. You may reflex from Custom Panel to Exome or Genome, or from Exome to Genome.

The User Filters allow filtering by variant tags assigned by the user or AI Shortlist:

• Most Likely – variants tagged as Most Likely by AI Shortlist or by the user.

• Candidates – variants tagged as Candidate by AI Shortlist or by the user.

• Incidental – variants tagged as Incidental by AI Shortlist or by the user.

• Carrier – variants tagged as Carrier by AI Shortlist or by the user.

• Not Relevant – variants manually tagged as Not Relevant.

• My Tags – variants tagged by the current user.

• AI Shortlist (Auto Analysis) - variants tagged by the AI Shortlist.

The collapsible Variant page sidebar allows you to access key case information, connect with Alamut and IGV, and view activity log, all within the Variant page. To expand the sidebar, click the arrow icon on the top right, and click again to collapse it.

The Variant page sidebar consists of three tabs:

1. Case Info tab displays relevant details about the case: sample names, the location of the BAM files connected to the case (if any), affected vs healthy sample status, and proband phenotypes.

2. Desktop App tab allows users to manage integration with the IGV and Alamut desktop applications.

3. Activity tab records variant-level user activities such as tagging a variant, adding comments, drafting variant interpretation notes, and editing the evidence graph. This aids collaboration and ensures a traceable record of variant interpretation.

The Emedgene Workbench offers a wide array of dynamic filters to help reveal or limit variants that are the most relevant to your clinical case. Each filter contains multiple options to customize the case review process according to your organization's best practices.

The Filters/Presets panel includes two tabs:

The Gene Filters allow filtering variants by genes or specific regions:

• All Disease Associated Genes - variants in the genes with a published disease association;

• All Unknown Genes - variants in the genes of unknown clinical significance;

• Candidate Genes - variants in a Gene list if defined during case creation;

• All ACMG genes - variants in the clinically actionable genes defined by the ACMG:
• Cancer Associated Genes - variants in genes with published association with oncological disease;

• LoF Genes (Emedgene Knowledgebase 26+) - variants in extremely LoF intolerant genes (gnomAD pLI ≥ 0.9). Note: if a variant is a CNV that overlaps more than one gene, it will appear in the filtering results if at least one of the genes has gnomAD pLI ≥ 0.9);

• Established HI/TS Genes (Emedgene Knowledgebase 26+) - variants in genes with sufficient evidence of dosage sensitivity (defined by having ClinGen's Haploinsufficiency and/or Triplosensitivity scores of 3);

• Coding regions - variants restricted to the protein-coding sequences.

Default values

The Variant activity panel on the righthand side of the Variant page records variant-level user activities, such as:

• Adding comments,

• Drafting Variant Interpretation notes,

• Editing evidence graph, etc.

The Inheritance Filters allow filtering variants using an assumed inheritance mode that is consistent with the segregation of genotypes and phenotypes in the family.

You can filter for:

• Autosomal Recessive - Homozygotes. Autosomal variants that are Hom in the Proband and other affected family members (if any) and Het, Ref, or No Cov in the unaffected members.

• Autosomal Recessive - Compound Heterozygotes. Two or more autosomal Het variants in the same gene inherited from different parents.

• X-Linked Recessive. X-chromosome variants in a male Proband (i.e. Hemi) that are Het in his unaffected mother.

• X-Linked Dominant. X-chromosome variants that are Het in affected females in the pedigree.

• De Novo Dominant. Autosomal and X-chromosome variants that are Het in the Proband and Ref in parents.

• Autosomal Dominant. Autosomal variants that are Het in the Proband and other affected family members but Ref in healthy relatives. ​

With the Display No Coverage slider, you can control whether to include or exclude variants that are No Cov in Proband or any other sequenced sample in that pedigree.

The Phenomatch Filters highlight variants located in the genes whose phenotypic annotation matches the proband's clinical presentation.

When one of the Phenomatch Filters is active, the results are limited to the variants that are top-rated by the corresponding algorithm:

• Phenomatch - High Specificity – strictly filters genes with disease phenotypes matching the exact patient HPO phenotypes.

• Phenomatch - Powered by AI – filters genes with disease phenotypes loosely matching the patient HPO phenotypes.

• Phenomatch with Unknown – filters genes of unknown significance based on indirect links to patient phenotypes (including mouse models, gene families, pathways, etc.).

Ethnicities of the proband's mother and father can be specified during the UI case creation. Starting from version 32.0.0, this can also be accomplished via API case creation. Please refer to the following list of supported ethnicities.

Case status indicates the current stage of case processing by the Emedgene platform or a genomic analyst.

Where can I find the current status of the case?

1. Status column of the Cases table;

1. Case details panel;

1. Individual case page Top bar.

Built-in statuses include:

1. Uploading

The sequencing file is being uploaded on the platform.

Note: The Uploading status cannot be manually altered.

2. In Progress

Case is running.

Note: The In Progress status cannot be manually altered.

3. Delivered

The running stage completed, and the case is now available for users for analysis and review.

4. Finalized

Analysis and review completed.

Status Finalized by design should be assigned manually after summarizing the analysis findings through the Case Interpretation widget. Specifically, when finalizing a case, you should indicate its end result:

• Confidently Solved (Positive),

• Likely Solved (Positive),

• Further Investigation (Uncertain), or

• Unsolved (Negative).

Confidently Solved, Likely Solved and Further Investigation end result categories correspond to the Resolved case status supercategory, and Unsolved obviously falls into Not resolved (together with all the non-finalized cases). The indicated case analysis outcomes are used to calculate the diagnostic yield.

5. Archived

Indicates that the case is not subject to further review. Changing Case status to Archived requires technical support (pre-34.0 flow).

6. Move to trash 🆕 34.0+

Makes the case unaccessible. This status is only assigned manually.

7. Pending Sequencing

The case has been created, but is not connected to any genetic data.

8. Issue Reported

The case failed to run. Please check the integrity of the files used and verify that the variant caller is on our accepted variant caller list.

9. Reanalysis

The system is re-running the AI Shortlist algorithm for the case.

How to change the Case status

A. On the Individual case page:

1. In the Individual case page Top bar, click on a dropdown icon next to the current case status.

2. Select the status from the menu.

B. In the Cases table:

1. In the Cases table, click on the current case status.

2. Select the relevant status from the dropdown menu.

Where can I manage Case statuses?

From Settings>Management>Test Statuses>Case Statuses, you can create custom Case statuses, remove unused Case statuses, and rearrange their order.

How to open a case

To open a case, mouse over the corresponding row in the Cases table and click on the Open case text next to the Case ID (first column). Alternatively, once a row is selected, clicking on it again will open the case as well.

How to filter cases

Go to Filters, select the field under Field, then choose or manually input value under List and click on Apply. To add another filter, click on Add new.

To remove a filter, under Active filters, click on the cross icon on the right of the filter setting. To clear filters altogether, click on the cross icon on the right of Filters.

Available filters include:

• Case Id

• Status: Issue reported, Uploading, In progress, Reanalysis, Archived, Delivered, Confirmed, Approved, Pending Sequencing, or any other customized case status

• Participants - users involved in the case submission, analysis, finalizing, or those who subscribed to receive updates on the case.

• Type: Custom Panel, Exome, Whole Genome

• Sample Id

• Resolved: Resolved, Not Resolved

• Label: any customized Case label

How to sort cases

You can sort cases by:

• Creation date, or

• Due Date.

To do this, hover over the column name and click on the up or down arrow to sort ascending or descending, respectively. The current sorting order is depicted as a single up or down arrow symbol next to the column's name.

Alternatively, you can click on the name of the column and select Sort ascending or Sort descending in the dropdown menu.

How to search for cases

You can use the Case search tab in the top bar to search for cases by the Case ID or Proband ID.

How to group cases

To group cases by Case Status, go to Group and select Status; to undo grouping, select None.

How to tweak Cases table view

1. Select fields to be displayed

2. Change column order

3. Change column width

Select fields to be displayed

A. Select fields to be displayed: go to Fields and set a toggle switch next to each field name in on or off position per your desired view.

B. Hide the currently displayed field: click on its title and select from the dropdown menu Hide column.

Change column order

A. Drag and drop columns: hover over the column title cell, click on the six-dot icon on the left to the text, drag to the desired location and drop.

B. Set column order under Fields: go to Fields, hover over the field name, click on the six-dot icon on the left to the text, drag to the desired location and drop.

C. Move a particular column: click on the column title and select from the dropdown menu Move left or Move right.

Change column width

Grab the right or left border of the column title cell and drag it with your cursor to the desired width.

​ ​

Add new case page > Case info screen > Select genes list

Select gene list

You can limit analysis to a gene list in the platform while creating a case. Choose between:

1. All genes

No limitation of the analysis.

2. Phenotype based genes

The list is automatically built from genes related to the HPO terms you entered for the case (per Emedgene knowledge base).

3. Existing gene list

Select one of the previously added gene lists from a dropdown list.

4. Create a new gene list

Generate a new virtual panel: add a List title and then add all the gene symbols one by one (Selection mode) or in a batch (Batch mode).

Selection mode

For each gene please follow the steps described below: Enter a gene symbol in the search box in the right panel (Candidate Genes) and select a matching symbol from a dropdown menu.

Batch mode

After selecting batch mode, paste a list of comma-separated gene symbols in the search box in the right panel (Candidate Genes).

Gene list modes

You can choose between two different modes of a gene list feature:

1. Gene panel mode

The default option.

Analysis is limited to the selected gene panel, no variants in other genes are considered in the results. If this in silico panel is used for analysis of exome or genome data, the gene restriction may be lifted during manual analysis to "open-up" the entire exome or genome for analysis.

2. Boost genes mode

Enabled through checkbox.

Analysis is performed for variants in all the genes. Variants in the targeted genes get upgraded scores during prioritization by the AI Shortlist algorithm.

How to filter variants by a gene list

1. Candidate Genes filter

To filter variants by the gene list, or remove the gene list restriction after the case has been delivered, go to the Analysis tools > Filters > Gene Filters > Candidate Genes filter.

2. Gene Lists filter created in a case

If you have run a case in All genes mode, you still can filter variants by a gene list. Go to Analysis tools > Presets > Gene Lists and select a list of interest. The Gene Lists filter presets can be defined by organization's manager.

3. Gene Lists filter created in Settings

Alternatively to creating a gene list filter in Presets, create and manage gene lists in Settings > Management > Gene lists.

First, fill in the List title and click Add button. Then add the gene symbols one by one (Selection mode) or in a batch (Batch mode).

• Selection mode - for each gene please follow the steps described below: Enter a gene symbol in the search box (Candidate Genes) and select a matching symbol from a dropdown menu.

• Batch mode: Paste a list of comma-separated gene symbols in the search box (Candidate Genes).

When you finished, press Save.

To view the Gene list in read-only mode, click on its name. By pressing the corresponding icons next to the Gene list's name, you can duplicate, edit or delete it. If a gene list has already been used by a case in the platform, you will only have the possibility to duplicate it.

To do this, click on the Export icon on top of the Analysis tools tab.

Log in to Emedgene and navigate to Settings in the upper right-hand corner of the page.

Click on the Management tab and then on Add Storage.

Choose Illumina BaseSpace storage type.

Fill Client Key, Client Secret and App Token as provided from BaseSpace (a description on how to get this information is provided below) and click Add storage to complete the setup.

• Via Command Line

• Via BaseSpace Developer Portal

Via Command Line

Prerequisite

Install BaseSpace CLI (Command Line Interface)

# Linux
$ wget "https://launch.basespace.illumina.com/CLI/latest/amd64-linux/bs" -O $HOME/bin/bs
# Mac
$ wget "https://launch.basespace.illumina.com/CLI/latest/amd64-osx/bs" -O $HOME/bin/bs
# or
$ brew tap basespace/basespace && brew install bs-cli
# Windows
$ wget "https://launch.basespace.illumina.com/CLI/latest/amd64-windows/bs.exe" -O bs.exe

Follow the instructions on the BaseSpace CLI Installation Page if needed.

Authenticate

On BSSH, login to the workgroup you want to connect as the storage.

Once the BaseSpace CLI is installed, run the authentication command in the terminal.

$ bs auth

The command will direct you to a link which requires to login.

After the authentication was completed successfully, find the access token in the config file.

$ cat .basespace/default.cfg

The result should look like -

apiServer   = https://api.basespace.illumina.com
accessToken = 

Populate the App_token with the accessToken value, and Server with the apiServer URL from the BSSH config file.

Client_key will be displayed in subsequent menus, so a descriptive name such as the workgroup name can be used.

Client_secret is unused when the App_token is available and can be set to "x".

Via BaseSpace Developer Portal

Go to the BaseSpace developer portal and login.

Go to My Apps and click Create a new Application.

Fill details for the application and click on create an application.

Fill details and press save.

You will need to fill all the fields that it requested, please add “NA” to them.

Go to My Apps and click on your new app. Then go to the credentials tab.

You will find the Client ID (Client Key), Client Secret and App Token to enter to Emedgene platform.\

Adding BSSH account to your Emedgene account

1. Log in into the desired Emedgene organization.

2. Go to Settings

3. Go to Management tab

4. Click on Add Storage

5. Select BaseSpace:

1. Add the information from your “Credentials” of the App previously created in BSSH.

When you:

• Complement NGS with other genetic tests done on the side (long-read sequencing, optical mapping, CGH, SNP array, karyotyping/FISH, repeat-primed PCR, MLPA, Southern blot, etc), or

• Choose to report a few adjacent variants as a single multi-nucleotide variant,

the need to add variants on top of the analysis results arises.

You can manually add variants absent from the VCF or not called from the FASTQ. Supported variant types are SNV, CNV, UPD, ROH, and STR. SV is coming soon!

To manually add a variant:

1. Click on the plus button on the top right of the Analysis tools tab.

1. In the Manually Add Variant window select variant type among SNV, CNV, UPD, ROH, and STR.

2. Fill in variant details according to the selected variant type:

1. Click on Create Variant.

Manually added variant's Variant page

Unlike regular variants, the manually added variant's Variant page has a blue frame and a "Manually added variant" title.

To filter by manually added variants:

In User filters select Manually added variants:

Add new case page > Family tree screen > Add patient information panel > Patient info section

1. Fill in the boxes:

1. Sex (33.0+) / Gender (≤32.0) (*)

Options: Male, Female, Unknown.

2. Relationship

Indicates the family relationship of a subject to the Proband automatically inferred from the pedigree. Options: Father, Mother, Sibling, Child, Other.

3. Date of Birth

Expected format: mm/dd/yyyy.

4. Ignore Sample

Mark the checkbox if you want to exclude the sample from the AI Shortlist analysis and Inheritance filters while preserving genotype data.

5. Add Proband's phenotypes

If a sample shares some phenotypes with the Proband, you can copy them by checking this box. Proband's phenotypes will appear in a newly created Related Phenotypes section. To remove any of the proband's phenotypes not observed in a current individual, click the ☒ button next to the HPO term in the Related Phenotypes section.

6. Unrelated Phenotypes

Phenotypes not shared with a Proband. They can be added one by one (Selection mode) or in batch (Batch mode).

Selection mode

Please follow the steps described below for each phenotype:

• Enter an HPO term (e.g., Hypoplasia of the ulna), an HPO ID (e.g., HP:0003022), or a descriptive phenotype name (e.g., Underdeveloped ulna) in the search box;

• Select a matching term from a dropdown menu and press Complete after you've added all the terms.

Batch mode

Paste a list of comma-separated HPO terms or HPO IDs (🆕 32.0+) in the search box and press Complete.

2. Click on Complete once all the information is added.

If you're comfortable with scripting and API usage, you can upload multiple cases at once using those methods. But if you're not a technical expert, don't worry. There is a user-friendly alternative available in versions 32.0 or newer - importing a CSV file directly through the user interface.

Please follow the steps as described below.

1. Prepare a CSV file

CSV (Comma-Separated Values) is a simple file format used to store data in tabular form. A row represents a sample, and a column represents a data field.

Start by downloading a CSV template with an example line and mandatory and non-mandatory fields from the Add new case page set to Batch mode (see step 2). Fill the file with your data according to CSV format requirements.

2. Upload a CSV file

1. Click on the + New case button on the Top navigation panel.

2. Click on the Switch to batch button in the top right corner. You'll be directed to the Select file page of the Batch upload flow. Note: Here you can download a CSV template in the valid format.

3. Drag and drop a CSV file into the box or upload it from the file explorer. Wait for file upload and validation to finish.

3. Review file validation results

After validation is complete, you will be directed to the Batch validation page. It features validation results details for you to review:

• File name,

• Number of rows in the file,

• Number of cases to be created

• Number of errors found,

• Status message:

  • if no errors were detected, a success message will be displayed;

* If any errors were detected, an error message will be displayed. You will be given the option to download a file with error details to help you diagnose and correct any issues with the data. Once you've corrected the CSV file, reupload it.

4. Create cases

1. Click on Create. A progress bar will appear on the right as the cases are created (Cases creation page).

2. If the cases have been created successfully, the Cases summary page will display the total number of cases that were created.

3. If there were any errors during the batch case creation process, the Cases summary page will display a table indicating the number of cases that were successfully created and the number of cases that failed.

You will have the option to download a CSV file containing two additional columns: Errors and Case ID. The Errors column will contain error messages for samples where case creation failed, while the Case ID column will contain the Case ID of a successfully created case for the lines where case creation was successful.

1) A Pathogenicity box

where you and other users in your group can manually assign variant pathogenicity by selecting an option from the dropdown (Pathogenic, Likely Pathogenic, VUS, Likely Benign, Benign).

If the variant is already in your Curate database, the previously selected pathogenicity will be marked with a Curate logo.

2) Variant Interpretation notes

that include basic variant details added by the AI Shortlist algorithm.

The notes can be manually edited (Edit text link). In editing mode, Paste icon becomes available. You can select actions from the dropdown menu, including:

1. Import data from Curate:

   1. Gene - import Interpretation from Curate Genes)

   2. Variant - import Interpretation from Curate Variants

2. Choose from related cases - connect summary notes available for the variant if it was classified in one of your organization's previous cases.

3. Choose from template - generate variant interpretation using the Variant interpretation template.

3) A See evidence button

under the Evidence box links to the Evidence page where you can assess more extensive evidence and generate an evidence graph for the variant under review.

4) ACMG SNV Classification wizard

to review and modify the automatically assigned sequence variant pathogenicity class. 23 out of 28 ACMG criteria are automated; the other five should be checked manually. On 32.0+. the software additionally calculates the variant score based on a points-based system recommended by the ACMG.

ACMG CNV Classification wizard

to review and modify the automatically assigned genomic variant pathogenicity class. Mostly automated.

5) Sanger Confirmation toggle button

to indicate if the variant should or has been submitted for validation through Sanger sequencing.

Note: Keep in mind that the Evidence section is active only for variants that have been automatically or manually tagged. To enable the Evidence section, you need to assign any tag to the variant under consideration.

The Variant page top bar displays:

1. Case ID,

2. Gene symbol,

3. Variant Type,

4. Genomic location of the variant,

5. Variant Interpretation notes if available,

6. Variant tag field.

   If the variant has been tagged manually (besides or instead of automatic tagging by the AI Shortlist), the user-selected tag will be shown. Otherwise, you'll see an automatically selected tag or N/A for no tag. Clicking on the Variant tag field lets you review a record of tagging in a dropdown, in the Assigned tags section. The variant tag menu also includes Viewed by section. Note: After a case reanalysis, all variants appear as not viewed.

1. A link to your Curate database.

V37 and below: If the variant is already in your Curate database, you will see an Open Curate button. Otherwise, you will see an Export to Curate button.

V38+: Clicking on this button will open a menu with 3 options:

1. If the variant is not already in your Curate database, you will see an Add to Curate button. Clicking this button adds the variant to Curate for further review and editing.

2. If the variant is already in your Curate database, and you have the “kms_export” role, you will see an Update in Curate button. Clicking this button updates the variant record in Curate with information from the current Analyze session, including:

   1. Pathogenicity

   2. Variant interpretation

   3. Gene-related disease (unless it is custom)

   4. Selected transcript (unless it is custom)

   5. ACMG data (classification, score, tags, tag strength, tag notes)

3. If the variant is already in your Curate database you will also see an Open in Curate button.

You can implement different combinations of Presets to be used for different case types (i.e. Presets for exome may be different from Presets for genome) as defined by your SOPs to further streamline case review.

The combination of Presets is referred to as a Preset group.

Select a Preset group to display in the case

Preset group selection is available in the Case info screen of the Add new case flow while creating or editing a case.

Where can I manage Preset groups? (34.0+)

To manage filter Preset groups, navigate to Settings > Organization Settings > Lab Workflow:

Preset groups

From here, you can create (from Presets/from a JSON file, edit, hide/unhide and download Preset groups as needed.

Default Preset group

Here, you can set a Preset group as default, so it will be used unless another Preset group is selected during case creation.

The Quality Filters allow one to filter variants by variant quality metrics. The filter can operate in a Simple or Advanced mode.

Modes

Simple

Select a minimum degree of sequencing Quality (Low, Moderate, High).

Advanced

For SNVs and Indels:

• Select a minimum degree of sequencing Quality (Low, Moderate, High),

• Define minimum Mapping Quality (0-60 - value can be exceeded using the text box),

• Specify minimum Depth (0-500 - value can be exceeded using the text box),

• Set minimum number of alternate reads in Alternate Read (0-500 - value can be exceeded using the text box). Note: available for cases run with pipeline version 5.26+.

• Set limits on Allele bias (0-100). Note: when applied to mtDNA variants, the Allele bias filter operates on heteroplasmy levels.

For CNVs:

• Set limits on CNV Length (50bp, 1kb, 10kb, 100kb, 1Mb, 100Mb, Max CNV length),

• Set minimum CNV Bin Count (1, 5, 10, 25, 50, 100, 500).

Default values

When Filters are Reset to Default, The Quality Filters are set to:

• Quality: Moderate and High,

• Mapping Quality ≥45,

• Depth ≥ 10,

• Alternate Read - no filtering,

• Allele bias - no filtering,

• CNV Length - no filtering,

• CNV Bin Count - no filtering.

As of version 2.26:

1. Fill in the boxes:

1. Sex (33.0+) / Gender (≤32.0) (*)

Options: Male, Female, Unknown.

2. Relationship

The default fixed value for Proband is Test Subject.

3. Date of Birth

Expected format: mm/dd/yyyy.

4. Medical Condition (*)

Options: Affected, Healthy.

The default value for Proband is Affected, but you may change it to Healthy.

5. Proband Phenotypes (*)

To add all relevant phenotypes for the Proband, use one of the following methods:

Selection mode

Please follow the steps described below for each phenotype:

1. Enter an HPO term (e.g., Hypoplasia of the ulna), an HPO ID (e.g., HP:0003022), or a descriptive phenotype name (e.g., Underdeveloped ulna) in the search box.

2. Select a matching term from a dropdown menu and press Complete after you've added all the terms and additional patient information below.

Batch mode

Paste a list of comma-separated HPO terms or HPO IDs (🆕32.0+) in the search box and press Complete.

Extract HPO terms from the file uploaded in the Clinical Notes section

In the Clinical Notes section upload a description of the clinical presentation in .pdf, .xls, .txt, .doc, .jpeg, or .jpg format. Among the extracted HPO terms for Phenotypes and Diseases select the ones you want to add to Proband's Phenotypes.

6. Proband Suspected Disease Condition.

Enter the disease name in the search box, select a matching term from a dropdown menu and press Complete. All the associated phenotypes will be automatically added to the Proband Phenotypes. To remove any phenotype described for the disease but not observed in your patient, click the ☒ button next to the HPO term in the Proband Phenotypes list.

7. Suspected Disease Penetrance

Enter the suspected disease penetrance as a percentage.

8. Suspected Disease Severity

Select the appropriate category to indicate the severity of the disease symptoms observed in the patient: Mild, Moderate, Severe, Profound.

9. Consanguinity

Mark the checkbox if applicable.

10. Patient Ethnicities

2. ​Click on Complete once all the information is added.

The finalized case status locks the case to prevent further changes to interpretation notes, ACMG tags, variant tags, pathogenicity and case-level interpretations.

How to finalize a case:

2. In the Case interpretation widget, indicate the final result of the analysis:

1. Confidently Solved (Positive),

2. Likely Solved (Positive),

3. Further Investigation (Uncertain), or

4. Unsolved (Negative).

The indicated case analysis outcomes are used to calculate the diagnostic yield.

You have the flexibility to reorder variants by drag-and-drop. The order of variants will be preserved in the Clinical Report within each variant table and/or section, defined by a variant tag and a variant type (e.g. SNVs tagged "In report").

Changes from v38.0+:

• Users will see a dropdown with an indication of number of variant that were assigned to each tag

  (eg. from screenshot - 'Most Likely (10)' )

• For Multi tag mode: A variant will show under all tags it is assigned to, along with the current selected state.

In Gene Interpretation, you can import gene annotation from Curate (30.0+).

5. To complete the Case interpretation flow, press Save.

You can download the report preview in a .pdf or .odt format.

All the generated reports are saved per case and each can be downloaded in a .pdf or .odt format.

Multiselection of variants:

1. Hover over a variant to reveal a checkbox at the start of the line;

2. Checking the box exposes the Multiselect actions bar which replaces the Search bar, and checkboxes appear for all variants in the current view;

3. Manually select variants one by one or check the Select all checkbox. Note: the Select all function applies solely to variants displayed on the current page, not all variants matching the active filters.

Bulk actions:

1. View/un-view variants

2. In the Multiselect actions bar, click on the Viewed icon and select Viewed or Un-viewed. Note: user-tagged variants can’t be un-viewed.

2. Assign/remove tags

How to assign a tag to multiple variants at once:

2. In the Multiselect actions bar, click on the Tag icon and choose a tag from the dropdown menu. Note: If any variants already possess tags, a warning message will appear. Click on Apply to proceed.

How to remove tags from multiple variants at once:

2. In the Multiselect actions bar, click on the Tag icon, then click on:

3. Assign/clear pathogenicity

How to assign pathogenicity to multiple variants at once:

2. In the Multiselect actions bar, click on the Pathogenicity icon and choose a pathogenicity class from the dropdown menu. Note: only tagged variants can have pathogenicity assigned.

How to clear pathogenicity from multiple variants at once:

2. In the Multiselect actions bar, click on the Pathogenicity icon, then click on Clear.

The Evidence page presents the most relevant evidence behind the automatic variant classification suggested by the AI Shortlist.

The Evidence page is accessible from:

Click on the variant bar.

View modes

You can switch between the graph and text view by clicking on the link on top of the page (Show evidence graph or Show evidence as text, respectively). The graph view is helpful for exploring the data, while the text view is relevant for collecting notes.

AI Shortlist collects data from credible studies and public databases in an internal knowledge base that maps complex connections between variants, genes, mechanisms, diseases, and phenotypes.

The Evidence page provides concise evidence for the specific variant, including:

Variant information

The main effect of the variant, its HGVS nomenclature for coding DNA and protein changes, and zygosity, including if the variant is de novo.

Gene information

Gene symbol, if the gene is tolerant to variation, and assumed inheritance mode in the case under review. This is suggested based on the observed level of genotype-phenotype co-segregation and inheritance mode of the genetic condition (reported or suspected).

In addition to the conventional inheritance modes (Autosomal Dominant, Autosomal Recessive, Compound Heterozygote Autosomal Recessive, X-Linked Dominant, X-Linked Recessive), the platform also employs Autosomal Dominant Partial Penetrance and Partial Autosomal Recessive designations.

Autosomal Dominant Partial Penetrance is used when the gene-associated condition is AD, and the variant is Het in the test subject and at least one of their parents. This suggests that the phenotypes may be due to incomplete penetrance of the genetic condition.

Partial Autosomal Recessive is used when the gene-associated condition is AR and the variant in the test subject is Het. This helps to account for the possibility that another causative variant is undetected - in the same (compound heterozygosity) or another (digenic inheritance) gene.

Disease information

Condition name as suggested by OMIM, other disease databases or in the literature.

Patient Phenotypes

Proband's phenotypes that match phenotypes reported for the suspected disease. Exact, indirect, and matches by ascendance are considered.

Unconfirmed Disease Phenotypes

Phenotypes reported for the suspected disease but not observed in the proband.

Unmatched Patient Phenotypes

Phenotypes observed in the proband but not known to be manifested as part of the suspected disease.

Links

Follow the links to the primary sources to explore the evidence further. The links are in the References section (text view) and are accessible by hovering over the arrows (graph view).

Edit mode

The evidence graph can be manually edited to include additional evidence for the case's resolution. To enter the edit mode, click on the pencil icon in the top left corner of the page. In this mode, you can edit, add, and delete text boxes.