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Tips & tricks. Christian Gilissen ( [email protected] ). What we do. Find genetic cause for rare Mendelian disorders Exome sequencing of patients / parents Find a single nucleotide change among thousands Only collaborative projects. How we do it. 3x SOLiD 4. 2x 5500xl.

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Tips tricks

Tips & tricks

Christian Gilissen

([email protected])


What we do

What we do

  • Find genetic cause for rare Mendelian disorders

  • Exome sequencing of patients / parents

  • Find a single nucleotide change among thousands

  • Only collaborative projects


How we do it

How we do it

3x SOLiD 4

2x 5500xl

50Mb Agilent SureSelect, 8 barcoded samples per run

About 20 samples per week

Completed >400 to date


How bioinformatics

How bioinformatics

  • 10 Node cluster running bioscope 1.3

  • Custom analysis pipeline

    • Annotation

    • Statistics & QC

    • Variant prioritization

    • “User friendly”

  • One for all!


How we do

How we do

  • Revisited projects from last year (100 exomes)

  • 10 projects with a dominant disorder

    • 6 new genes identified

    • Found mutations in 3 known genes

  • 14 projects with a recessive disorder

    • 8 new genes identified

  • A success rate of ±60%


Why did we fail

Why did we fail?

  • Gene is not in the enrichment kit

  • Lack of sequence coverage

  • Bioinformatic variant calling issues

  • Bioinformatic annotation issues

  • Mutation is not within the coding sequences.

  • Clinical heterogeneity or wrong diagnosis

  • Not a SNV

  • Others publish first


Tips tricks exome sequencing

Tips & Tricks exome sequencing

  • Better be too strict than too relaxed

  • Reanalyze your data

  • Focus your efforts where it matters most

  • Make your life easy: automate, keep uniformity

  • If at first you don’t succeed, use your common sense

  • Be independent


Current bioinformatics setup

Current bioinformatics setup

  • Common programming language

  • IDE: Eclipse

  • Code repository

  • Ticket system

  • Automated build scripts

  • Automated testing

  • Release procedures

  • Common coding framework


Bioinformatics

Bioinformatics

  • All bioinformaticians in a single room

  • Shared framework

  • Write easy reusable code

  • Good collaboration with lab

  • “Protected” from individual collaborators


New developments

New developments

  • 5500xl instrument + lifescope

  • Pooled samples

  • Different enrichments

  • RNASequencing

  • IonTorrent

  • Diagnostics


Acknowledgements

Acknowledgements

Next generation sequencing team -

Genomic Disorders Group:

Bioinformatics

Jayne Hehir-Kwa

Joep de Ligt

Nienke Wieskamp

Marisol del Rosario

Eugene Verwiel

Lab

Alexander Hoischen

Lisenka Vissers,

Kornelia Neveling

Peer Arts, Bart van Lier, Marloes Steehouwer, Petra de Vries, Irene Janssen,

Joris A Veltman

http://genomicdisorders.nl


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