Exploring targeted long-read sequencing technologies to validate genetically engineered alleles generated by CRISPR technologies

Deadline for applications has passed.

Key information

Applications closed
05 October 2023, 11:59 BST
Hours per week
36 (full time)
Application guidance
Posted 30 August 2023
Background texture taken from the lab imagery.

This sandwich placement will be based in the Advanced Sequencing Facilities STP & Genetic Modification Service STP supervised by Jerome Nicod and Ben Davies.

Project background and description 

With the advent of CRISPR and other genome engineering technologies, our ability to manipulate the mouse genome has increased, allowing more accurate ways to model human genetic disease. At the Crick, there is a growing demand for mouse models involving complex and large-scale genome engineering. As the complexity of the mutations increases, it’s crucial to implement enhanced methods for the validation and quality control of these engineered alleles. Short-read  sequencing is not always suitable, as it fails to detect common aberrations associated with CRISPR, such as tandem duplications. In contrast, long-read sequencing technologies overcome this limitation and enable the phasing of multiple mutations to be verified on a single allele.

In order to facilitate the verification/QC of the edited genome, it is advantageous to focus the sequencing to the target gene. Two technologies are available for this purpose: CRISPR-Capture and adaptive sampling. The former uses the CRISPR/Cas9 site-specific nuclease itself to cleave out the target locus from the genome, enabling its enrichment and sequencing; while the latter involves real-time alignment of the raw reads to a reference, enabling enrichment of a sequence of interest.

The student will test these two selective sequencing technologies on various engineered alleles, comparing results against untargeted whole-genome sequencing. The relative advantages of both technologies will be assessed in terms of cost, accuracy, ease-of-use and speed. The overall aim of the project is to design the best validation method, and implement it as standard workflow for all GeMS generated mutants at the Crick. This will have a direct impact on quality of alleles used by Crick researchers.

Candidate background

The post holder should embody and demonstrate the Crick ethos and ways of working: bold, open and collegial. The candidate must be registered at a UK Higher Education Institution, studying in the UK and must have completed a minimum of two years’ undergraduate study in a relevant discipline, and on track to receive a final degree grade of 2:1 or 1. In addition, they should be able demonstrate the following experience and key competencies:

  • The project would suit a biomedical student, with enthusiasm to gain experience in cutting edge technologies. The project has potential for a large bioinformatic component, so a student with computational skills would be advantageous, although full training will be given.
  • Good knowledge in relevant scientific area(s)
  • Good written and spoken communication skills
  • Ability to work independently and also capable of interacting within a group

References

1.         Gilpatrick, T., Lee, I., Graham, J.E., Raimondeau, E., Bowen, R., Heron, A., . . . Timp, W. (2020)

            Targeted nanopore sequencing with Cas9-guided adapter ligation.

            Nature Biotechnology 38: 433-438. PubMed abstract

2.         Payne, A., Holmes, N., Clarke, T., Munro, R., Debebe, B.J. and Loose, M. (2021)

            Readfish enables targeted nanopore sequencing of gigabase-sized genomes.

            Nature Biotechnology 39: 442-450. PubMed abstract