Name and Surname of the Supervisor: Giancarlo Russo
Institution (Company), Department: EMBL PI
Preliminary Topic: Optimising sgRNAs for HIF targets in PDAC via an in-vitro Cas9 cleavage assay and generating stable knockouts in PDAC cells
Short Description of the Internship Work: As part of a broader effort to study hypoxia pathways in pancreatic ductal adenocarcinoma (PDAC) cell lines, this MSc project benchmarks the in-vitro cleavage efficiency of sgRNAs targeting HIF1A, EPAS1 (HIF2A), and HIF3A to inform later single-, double-, and triple-knockout designs. The student will generate sgRNAs by designing and cloning guide sequences into T7-promoter plasmid templates, transcribing them in vitro, and assembling Cas9-sgRNA RNPs. Candidate sgRNAs - selected from the literature and design tools - will be assayed on PCR amplicons spanning each target site in single-tube reactions and quantified by gel and automated electrophoresis to compute the fraction cleaved versus uncut. In addition, the student will select the most efficient sgRNAs and use them to generate stable knockout PDAC cell lines with the MuLE lentiviral system by cloning single- and multi-sgRNA cassettes, producing lentivirus, transducing cells, selecting edited pools, and validating edits (and, time permitting, protein-level or qPCR readouts under hypoxia). Deliverables include a ranked sgRNA list per gene, recommended single-, double-, or triple-guide sets for follow-up, and at least one validated stable knockout line (or pool) ready for downstream hypoxia studies.
Qualification Requirements (Degree): MSc
Application Deadline: 2026-01-15
Additional Comments:
Supervisor: Giancarlo Russo
Consultant: Rūta Matulevičiūtė
Language: This project and the MSc thesis are conducted in English (lab communication & documentation in English).