Posters and guidelines
Thank you for considering to present your work as a poster at Oligo 2020 Oxford Virtual.
Digital poster submission deadline: Prepare your poster as you would normally do for printing, and submit your final poster as both PDF and JPG/PNG files via the link below no later than 26th March 2021. Late posters may not be included in the conference programme. Please DO NOT send your poster files by email.
Naming your poster files: Name your poster files as follows: <your surname>-GEOX21V-Poster.pdf | <your surname>-GEOX21V-Poster.png | <your surname>-GEOX21V-Poster.jpg, etc. For example, for David Jones, name your file as Jones-GEOX21V-Poster.pdf. DO NOT name your poster files as, e.g., Oxford-poster, poster2021v, Oxford-genome-editing-poster. Such files will be automatically rejected.
Poster presentation: Posters will be made available via a secure page to the symposium participants before the meeting. There will be two ways to interact with the poster presenters:
- the participants will be able to ask questions via the Zoom chatbox during the mid-conference break ; and/or
- the participants can post their questions on Twitter at any time using the meeting hashtag #GEOx21V, as well as the poster specific hashtag (given under each poster abstract) – do tag @LPMHealthcare in your tweets.
The poster presenters should regularly check Twitter for any questions about their posters before, during and after the meeting, and post their answers on Twitter using appropriate hashtags, as above.
Any further information about the poster presentations at this digital meeting will be available in the future.
Before uploading your poster, you must make sure that you follow ALL of the instructions above!
(Presenters in Bold)
Accepted poster abstracts (Unedited) will be published below. If your abstract has been accepted for presentation but it does not appear in the list below, please let us know as soon as possible by emailing CRISPROxford@gmail.com.
A robust pipeline for high throughput CRISPR/Cas-mediated manufacture of engineered knock-out and knock-in cell lines
Hashtags: #GEOx21V, #DBlakemore
Daniel Blakemore, Simon Pollack, Victor van Gelder, Hind Ghezraoui, Sonia Moratinos, Jack Williams, Holly Prangley, Yves Du Toit, Claire Greenwood, Matt Burridge, Pela Derizioti, Ryan Cawood, Suzanne Snellenberg
OXGENE, Medawar Centre, The Oxford Science Park, 1 Robert Robinson Ave, Oxford OX4 4HG, UK
The capacity to induce genetic modification in mammalian cells using CRISPR/Cas technology has the potential to revolutionise how we understand gene function. However, high-throughput application of this technology remains challenging, especially for complex alterations that rely on sporadic activity of the homology directed repair (HDR) pathway. In addition, cell types such as iPSCs are sensitive to these procedures, making it difficult to harness their potential. Here we present an automated, streamlined pipeline for generation of simple and complex gene modifications in multiple cell types including iPSCs. We first optimise transfection conditions, recovery from single cell sorting of the host cell lines and determine copy numbers of genes to be edited. In-house software, GRNADE and PRIMAPE, enables optimal sgRNA and primer design. We use ssODNs, the preferred template for HDR applications, for precise genome edits, and ascertain on-target cleavage and HDR efficiencies using in-house CRITIC software. We single cell sort transfected cells and use CellMetric® imaging software for automated and systematic monitoring to assure single cell clonality. Hamilton-robotics facilitate automated clone transfer for effective expansion. Clone screening for the correct edit is performed by CRITIC, followed by NGS analysis for validation. Using this pipeline, we have successfully generated knockouts and knock-ins in over 25 different cell lines, including iPSCs, for multiple applications from reagent verification and basic biology studies to complex disease modelling. We routinely achieve >95% editing efficiencies and up to 30% HDR efficiency using our optimised conditions. Also, the pipeline is optimised for multiple well-known iPSC culture systems, allowing us to meet demand for custom iPSC manufacture. Coupling CRISPR/Cas technology with high-throughput robotics facilitates scaling of simple complex genetic modifications, enabling large scale exploration of disease models.
Enhancing SaCas9 target specificity by rational directed mutagenesis
Hashtags: #GEOx21V, #HGhezraoui
Hind Ghezraoui, Gloria Mesa-Gil, Richard Parker-Manuel, Ryan Cawood, and Suzanne Snellenberg
OXGENE, Medawar Centre, Oxford Science Park Oxford, OX4 4HG, United Kingdom
Continued development of the CRISPR/Cas9 system is making therapeutic gene editing a viable biomedical tool. However, delivery of Streptococcus pyogenes (SpCas9) by adeno-associated viral vectors (AAV) is challenging due to the small packaging capacity of AAV (∼4.7 kb). This can be overcome by using the minimal SpCas9 ortholog Staphylococcus aureus Cas9 (SaCas9). Unfortunately, while many high-fidelity SpCas9 variants have been reported, far fewer SaCas9 variants have eliminated off-target editing, leaving off-target cleavage of unintended genomic sites a critical issue to be resolved for this ortholog. We developed a reporter plasmid that can be used to examine on- and off-target editing efficiency and used it to screen rationally engineered SaCas9 variants to identify those SaCas9 variants with enhanced specificity that could effectively discriminate a single base mismatch. We identified SaCas9 variants that dramatically reduced off-target effects, whilst maintaining robust on-target activity comparable to wild-type SaCas9. We confirmed editing at three genomic loci by CRITIC (CRISPR EdiTing InterferenCes) analysis using Sanger trace sequences. Thus, our variants could be used for genome editing applications where high fidelity is required.
Understanding the resistance landscape to gene drives targeting the doublesex gene in the malaria mosquito
Hashtags: #GEOx21V, #IMorianou
Ioanna Morianou1, Andrew M. Hammond1,2, Tony Nolan1,3, Andrea Crisanti1
1Department of Life Sciences, Imperial College London, London, UK
2Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Johns Hopkins University, Baltimore, USA
3Department of Vector Biology, Liverpool School of Tropical Medicine, Liverpool, UK
Gene drives are selfish genetic elements with the potential to spread throughout entire insect populations for sustainable vector control. Recently, a CRISPR/Cas9-based gene drive was shown to eliminate caged populations of the malaria mosquito by targeting the female-specific exon of the highly conserved doublesex gene. However, the key question remains whether natural populations might be able to evolve resistance to the gene drive. Resistant alleles may be naturally occurring or generated by the drive itself. To investigate the potential for resistance at doublesex, we developed a high-throughput mutagenesis screen designed to enhance end-joining mutations at the gene drive target site. We discovered a few putatively resistant mutations that get generated at a very low frequency. This strategy can be used to evaluate gene drives for potential resistance prior to field testing. Informed by this assay, we developed a third generation gene drive design that can mitigate against known resistance and potentially eliminate malaria from entire regions in Africa.