Forward genetic screens in haploid mammalian stem cells: looking beyond CRISPR/Cas9
Recent adaptation of the CRISPR/Cas9 bacterial system to facilitate manipulation of mammalian genomes has provided a real breakthrough for genome editing applications. Development of whole-genome CRISPR libraries with the aim of generating gene knockouts for every single coding sequence has allowed forward genetic screening in mammalian cells with unprecedented efficiency and versatility. CRISPR/Cas9 approaches, however, rely on phenotypes associated with loss-of-function mutations. Single-nucleotide variations (SNVs), on the other hand, have the potential to uncover not only loss-of-function phenotypes (by generating nonsense mutations, for example) but also gain-of-function phenotypes through missense mutations. In addition, they produce valuable information regarding functionally important domains of the affected gene product, as SNVs causative of a particular phenotype tend to cluster around specific regions of the amino acid sequence of the encoded protein. SNV-based approaches in mammalian cells, however, have been hindered by the diploid nature of their genomes, a fact that complicates the establishment of straightforward genotype-to-phenotype correlations. In this talk I will discuss how we apply CRISPR/Cas9 genetic screening to further understand the DNA-damage response in mouse embryonic stem cells (mESCs), and I will also introduce the use of haploid mESCs to perform SNV-based forward genetic screens.
Date:
16 July 2015, 14:30 (Thursday, 12th week, Trinity 2015)
Venue:
MRC Weatherall Institute of Molecular Medicine, Headington OX3 9DS
Venue Details:
Seminar Room
Speaker:
Dr Josep Forment (University of Cambridge)
Organising department:
MRC Weatherall Institute of Molecular Medicine
Organiser:
Penny Berry (University of Oxford, Department of Oncology, Weatherall Institute of Molecular Medicine)
Organiser contact email address:
penny.berry@oncology.ox.ac.uk
Host:
Dr Peter McHugh (University of Oxford, Weatherall Institute of Molecular Medicine)
Part of:
WIMM Occasional Seminars
Booking required?:
Not required
Audience:
Members of the University only
Editor:
Penny Berry
