SARS-COV2 SEQUENCING

ABOUT

 

With the advent of the covid-19 pandemic, the need arose for genomic surveillance to better understand the epidemiology, identify clusters and investigate outbreaks. To accomplish this, a partnership with IBC-ULB (Institut de Biologie Clinique ULB), ULB Erasme and the microbiology unit of UZ Brussel was initiated.

Since July 2021, BRIGHTcore has been sequencing Sars-Cov2 samples for diagnostic and research purposes.

METHODOLOGY

 

RNA is extracted from naso- and oropharyngeal swabs from Covid-positive patients (e.g. confirmed through qPCR). Samples, preferentially with a viral load > 3 log copies/ml are retained for sequencing.

The extracted RNA is converted to cDNA with a reverse transcriptase (NEB LunaScript) and subsequently used as input for a multiplex PCR (IDT Midnight PCR panel), to specifically enrich for the viral genome and reduce the human transcriptome background.

Next, the multiplex PCR amplicons are processed through the Oxford NanoPore Technologies Rapid Barcoding kit 96 (SQK-RBK110.96) and sequenced on the GridION x5 using a maximum of 96 samples per flow cell.

The full protocol from Nikki Freed et al. can be consulted here : full protocol. We slightly modified this protocol with additional (degenerate) primers as to compensate for PCR dropouts caused by the large degree of variations in the Omicron lineage.

More information on WGS and MPS is available through the buttons below.

 

INSTRUMENTS

The following devices are used for Sars-Cov2 sequencing in our premises.

NanoPore GridION x5

Sequencing long reads is no hassle on this device. It can sequence 5 MinION flowcells simultaneously and perform the data analysis on board.

 

CERTIFICATES

Sciensano recognized
Sciensano recognized

DOWNLOADS

 
Flyer2 GW testing white borders_edited.j
Sars-Cov2 SNV stats - PNG

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Flyer2 GW testing white borders_edited.j
Verzendinstructies Sars-Cov2 - NL

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Flyer2 GW testing white borders_edited.j
Sars-Cov2 consensus fasta - fasta

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Flyer2 GW testing white borders_edited.j
Sars-Cov2 coverage stats - PNG

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Flyer2 GW testing white borders_edited.j
Sars-Cov2 summary file - csv

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CONTACTS

 
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BRIGHTcore general inquires

+32 (0)2 477 64 79 (sec)

UZ Brussel

BRIGHTcore

Laarbeeklaan 101

1090 Brussels

pas14klein.jpg

Toon Janssen

+32 (0)2 477 64 79 (sec)

UZ Brussel

BRIGHTcore

Laarbeeklaan 101

1090 Brussels

pas14klein.jpg

Ben Caljon

+32 (0)2 477 64 79 (sec)

UZ Brussel

BRIGHTcore

Laarbeeklaan 101

1090 Brussels

SAMPLE TYPES

 

We aim to keep te list with sample types updated. However, if you believe we offer this test on other sample types, of if you have a very specific sample type you'd like to evaluate : please contact the corresponding persons for either diagnostic or research purposes.

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Tissue
Recipient
Quantity
Transport
Purpose / Extract
Sars-Cov2 RNA (naso/oropharyngeal
Microtube* or 96 wel plate (plate is preferred)
Log(copies/ml) > 3 (min. 5 µl)
Ice (max. 20 min) or dry ice
RNA

* Microtube can be  a either a cryovial or eppendorf tube (0,5 ; 1,5 or 2 ml). Please consider the quality of the tube used (nuclease free ; free of RNA/DNA ; sterile ; LoBind) according to application needed.  

 

DELIVERABLES

Below, you can find the type of data files that can be retrieved for this test.

If you require more information or need a more custom output, please consult our Bioinformatics page.

Deliverable
Description
Research or diagnostic
Sars-Cov2 consensus fasta
Consensus Sars-Cov2 genome (fasta format) for all samples in run/centre
Diagnostic / RUO
Sars-Cov2 SNV stats
PNG file to visualize the quality of the individual SNVs on the report
Diagnostic / RUO
Sars-Cov2 coverage stats
PNG file to visualize the completeness of Sars-Cov2 genome coverage
Diagnostic / RUO
Sars-Cov2 summary file - csv
Comprehensive report with quality metrics and genotyping results
Diagnostic / RUO
Raw data - fast5 file
Raw data file generated by Oxford NanoPore sequencers
RUO (on request)
Annotated variant file
Text file containing all detected variants + relevant annotation (cross reference to various databases)
Diagnostic / RUO
 

PLAN EXPERIMENT

1. For diagnostics

The decision to determine/sequence the full Sars-Cov2 genome is up to the microbiology or covid-lab, as this is only allowed for in specific cases/indications. As this is a diagnostic assay, the results are handled through your General Practitioner.

2. For Research

Alternatively, you may want to set up a study, e.g. to evaluate certain clusters, to assess immune evation (efficacy of vaccines),... For this, please get clearance from your local ethical committee and/or register your samples with a biobank

To set experiments up with us, please follow the steps below.

Get in touch (ONLY FOR NEW PROJECTS)

Explain your project, so we can assist you to find the best solution.

Expect questions like :

  1. What type of sample would you like to analyze?

  2. Will you handle the RNA extraction (please know : BRIGHTcore doesn't handle human pathogens)?

  3. In which elution buffer did you elute your RNA (we prefer nuclease free water)?

  4. How many samples would you like to process?

  5. How fast do you need the results (TAT) and/or do we need to expedite?

  6. If you deviate from our sample types, are there test samples available?

  7. Do you require bioinformatics support or are our standard deliverables ok?

  8. ...

Call Center Headset

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Request quotation

Once you know which experiment you want to set up, fill out the 'Quote request' form, with the essential details of your project :

  1. Coordinates of the person to whom to address the quotation

  2. Which type of test you'd like to set up

  3. Do you require bioinformatics support?

  4. Do we need to expedite (comes at extra cost)?

  5. ...

Image by Andre Taissin

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Submit your samples

Once you agree with our terms (quote / TAT / terms & conditions)  :

  1. Fill out our sample submission form

    1. Send it via email : contact us

    2. Print out the form and include it with your samples 

  2. Send your samples to us

    1. Adhere to the corresponding transport conditions listed under Sample types

    2. Label your tubes/plates correctly : see details here

    3. Please attach the 'verzendinstructies' (see downloads) on the shipping container (for RNA)

  3. Indicate if we can discard your samples​ after completion of project

Blood Samples

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Your experiment starts

Now it is up to us... We will start your experiment as soon as possible. When expedited, we will start the experiment within 1 week​.​ For standard requests the turn-around-time (TAT) varies :

  1. In diagnostics :  5 workdays

  2. In research : 4 weeks (or otherwise agreed)

The research TAT is generally overestimated, but this extended timeframe allows us to : combine similar requests of multiple scientists and to repeat the experiment in case of issues. 

Please refrain from contacting us in case the pre-agreed TAT didn't expire yet.

Alarm Clock

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