Next Generation Sequencing

Next generation sequencing technology has now reached tremendous throughput and accuracy that enable researchers to conduct large-scale genomic projects at affordable cost and in less time.

The NGS Platform of the Institut Curie has greatly evolved since 2012 mainly with the financial support of the ICGEx grant obtained in the frame of the EQUIPEX « investissement d’avenir » piloted by the ANR (ANR-10-EQPX-03). This grant allowed to purchase high throughput NGS sequencers, to fully equip a dedicated lab at the 7th floor of the hospital and expand the staff in charge of running sequencing projects.

 Aims

  • Provide state of the art services in sequencing for the different research teams of the Institut Curie.
  • Offer technical and scientific support in experimental design and sequencing strategies.
  • Be flexible to best respond to scientific needs (technical development and implementation).

Activity

Under the supervision of Olivier Delattre and Alain Nicolas, the NGS platform provides state of the art services in sequencing. This includes the organization of pre-run meetings where the choice of the most appropriate technical strategy is defined in order to best address the biological question of collaborators. Multiple sequencing protocols have been developed and routinely used at the platform in order to cover most commonly used applications in genetics and epigenetics (Genome, Exome and targeted resequencing, RNA sequencing, ChIP-seq …). Furthermore, we are also willing to co-develop more specific protocols according to the need of research teams of the Institute.

Once the technical aspects are defined, samples are provided by collaborators and libraries compatible with the dedicated sequencing devices (Illumina or PacBio) are then prepared by the NGS team. After sequencing, raw data are transferred to the bioinformatics team (U900) for quality controls and basic analysis. At the end, sequence data are provided to research teams with comprehensive analysis and quality reports.

The platform is not restricted to fundamental research, we are also involved in multiple translational research projects in collaboration with U830, the translational research department and the Curie Hospital.

Networking

  • Active member of the national infrastructure France Génomique, a French consortium of NGS platforms dedicated to the mutualisation of NGS resources that benefit to French research teams
  • Member of the European Core For Life network, which brings together several platforms of different technologies. The Core for Life Genomics Work Group provides networking opportunities enabling the sharing of experiences and knowledge.

Training

  • Organisation of the genomic module for the Master 2 degree « Ingénieur de plateforme » of Université de Paris.
  • Oral presentation on NGS for the Licence Pro Génomique of ENCPB (Paris).
  • Oral presentation in « International Courses » organized by the training unit of the Institute.
  • Oral presentation on NGS for the EPF Engineering school (Sceaux).
  • Oral presentation in Bioinformatic training organized by the U900.
  • Oral presentation on NGS for the Master 2 degree of the « Ecole doctorale de Cancérologie de Paris » (Saclay).

Services

  • Availability of a dedicated interface for NGS projects submission and follow up
  • Library preparations for multiple sequencing applications :
    • Whole genome, whole exome and targeted resequencing
    • Transcriptome analysis (RNAseq for mRNA and total RNA)
    • Epigenetics with ChIP-seq for histone modifications or protein binding analysis. Analysis of open regions of chromatin from single cell (single-cell ATAC-seq)
    • Single cell mRNA sequencing (3’ seq technology and Spatial Transcriptomic of 10X Genomics)
  • Development of analysis pipelines for routine applications
  • Bioinformatics supports through collaboration with the bioinformatics platform (U900)

 Equipment

  • One high throughput sequencer from Illumina : 1 NovaSeq 6000 dedicated to applications requiring a large amount of data (human genome and exome sequencing, transcriptome analysis …).
  • Two benchtop sequencers from Illumina : 2 MiSeq used for projects requiring a low depth of sequencing and for custom libraries validation.
  • One Long Read sequencer from PacBio : 1 Sequel II allowing the sequencing of long fragments (>10kb) and repeated regions.
  • One Chromium from 10X Genomics enabling single cell analysis, including transcriptomics.
  • 1 LabChip GXII Touch, 1 BioAnalyzer, 1 Femto Pulse, 1 CFX96 real-time PCR system et 1 Pippin Pulse used to assess samples and libraries quality.

All equipments have been funded by ICGex. Except the Novaseq cofunded by ITMO Cancer and Hôpital Curie. The Chromium from 10X Genomics has been funded by SiRIC.

PacBio Sequel II Long read sequencer
PacBio Sequel II Long read sequencer
Illumina NovaSeq 6000 high-throughput sequencer
Illumina NovaSeq 6000 high-throughput sequencer

To contact us :

Next actions for 2021

  • Implementation of the Visium Spatial Gene Expression technology from 10X Genomics.
  • Implémentation of the HiFi reads application with the Sequel II from Pacific Biosciences (reads with >Q20 quality and several kb length).
  • Setting-up of a Quality approach in order to obtain ISO 9001 certification.

Key publications

Year of publication 2020

Sophie Loeillet, Mareike Herzog, Fabio Puddu, Patricia Legoix, Sylvain Baulande, Stephen P. Jackson, Alain G. Nicolas (2020 Oct 6)

Trajectory and Uniqueness of Mutational Signatures in Yeast Mutators

Proceedings of the National Academy of Sciences : 117 n° 40 : 24947-24956 : DOI : 10.1073/pnas.2011332117
Yann Kieffer, Hocine R. Hocine, Géraldine Gentric, Floriane Pelon, Charles Bernard, Brigitte Bourachot, Sonia Lameiras, Luca Albergante, Claire Bonneau, Alice Guyard, Karin Tarte, Andrei Zinovyev, Sylvain Baulande, Gerard Zalcman, Anne Vincent-Salomon, and Fatima Mechta-Grigoriou (2020 May 20)

Single-Cell Analysis Reveals Fibroblast Clusters Linked to Immunotherapy Resistance in Cancer

Cancer Discovery : DOI : 10.1158/2159-8290.CD-19-1384
Marie-Ming Aynaud, Olivier Mirabeau, Nadege Gruel, Sandrine Grossetête, Valentina Boeva, Simon Durand, Didier Surdez, Olivier Saulnier, Sakina Zaïdi, Svetlana Gribkova, Aziz Fouché, Ulykbek Kairov, Virginie Raynal, Franck Tirode, Thomas G P Grünewald, Mylene Bohec, Sylvain Baulande, Isabelle Janoueix-Lerosey, Jean-Philippe Vert, Emmanuel Barillot, Olivier Delattre, Andrei Zinovyev (2020 Feb 13)

Transcriptional Programs Define Intratumoral Heterogeneity of Ewing Sarcoma at Single-Cell Resolution.

Cell reports : 30 : 1767-1779.e6 : DOI : 10.1016/j.celrep.2020.01.049
Floriane Pelon, Brigitte Bourachot, Yann Kieffer, Ilaria Magagna, Fanny Mermet-Meillon, Ana Costa, Anne-Marie Givel, Youmna Attieh, Jorge Barbazan, Laetitia Fuhrmann, Stéphanie Descroix, Danijela Vignjevic, Pascal Silberzan, Isabelle Bonnet, Claire Bonneau, Maria Carla Parrini, Anne Vincent-Salomon & Fatima Mechta-Grigoriou (2020 Jan 21)

Cancer-associated fibroblast heterogeneity in axillary lymph nodes drives metastases in breast cancer through complementary mechanisms

Nature Communication : 11 : 1-20 : DOI : 10.1038/s41467-019-14134-w
Rafael Galupa, Elphège Pierre Nora, Rebecca Worsley-Hunt, Christel Picard, Chris Gard, Joke Gerardavan Bemmel, Nicolas Servant, Yinxiu Zhan, Fatima El Marjou, Colin Johanneau, Patricia Diabangouaya, Agnès Le Saux, Sonia Lameiras, Juliana Pipoli da Fonseca, Friedemann Loos, Joost Gribnau, Sylvain Baulande, Uwe Ohler, Luca Giorgetti, Edith Heard (2020 Jan 16)

A Conserved Noncoding Locus Regulates Random Monoallelic Xist Expression across a Topological Boundary

Molecular Cell : 77 n°2 : 352-367.e8 : DOI : 10.1016/j.molcel.2019.10.030

Year of publication 2019

Roberta Ragazzini, Raquel Pérez-Palacios, Irem H Baymaz, Seynabou Diop, Katia Ancelin, Dina Zielinski, Audrey Michaud, Maëlle Givelet, Mate Borsos, Setareh Aflaki, Patricia Legoix, Pascal W T C Jansen, Nicolas Servant, Maria-Elena Torres-Padilla, Deborah Bourc'his, Pierre Fouchet, Michiel Vermeulen, Raphaël Margueron (2019 Aug 26)

EZHIP constrains Polycomb Repressive Complex 2 activity in germ cells.

Nature communications : 10 : 1-18 : DOI : 10.1038/s41467-019-11800-x
All publications