Project

Developing a hiPSC reporter cell line for cardiac development

Publisher

Supervisor

Location

Greater Copenhagen area

Master’s project, 60 ECTS, start in spring 2019

Bioneer A/S and DTU Food

Aim:

This project aims to develop a hiPSC reporter cell line for cardiac development by introducing the luciferase gene with CRISPR/Cas9. Luciferase expression during cardiac development will be measured, and ultimately validated for use as a predictive toxicity assay.

Background:

Human induced pluripotent stem cells (hiPSC) hold great potential for assays within predictive toxicity testing, drug development and disease modeling. They are human cells and therefore model the human biology best, although, in contrast to laboratory animals, they cannot recapitulate a whole organism. However, hiPSC can mimic certain tissues, organs, and cellular mechanisms. This is particularly needed in research fields where good alternative models are needed, such as in developmental toxicity. Developmental toxicity focuses on the developing embryo and fetus, and tests the hazards of chemicals to the embryo or fetus via exposure through the mother. Because embryonic development is a complex process in humans compared to laboratory animals, this is particularly challenging to test with animals.

Human embryonic development can be modelled using hiPSC that form so called embryoid bodies (EBs) that are structurally and functionally in many ways similar to human embryos. EBs can generate most cell types of the embryo, but if guided appropriately, they can also be differentiated into only one cell type, as for example cardiac cells of the heart. This differentiation into cardiac cells has previously been used in a developmental toxicity test (the embryonic stem cell test, EST) using murine or human embryonic stem cells. However, using hiPSC no ethical issues arise, and furthermore the readout has previously been based on microscopy analysis, which is limited in its throughput.

We developed a novel EST-like test based on hiPSC in a 96-well plate format that can be used to screen chemicals for developmental toxicity. Now we are aiming to develop a simplified read-out method, based on measuring luminescence, which is suitable for high-throughput screening. We plan to introduce the luciferase gene by using CRISPR/Cas9, a recently developed method for targeted gene editing in human cells. The developed reporter cell line will be used to generate EBs and to differentiate into cardiac cells, with luciferase expression as the readout. The final aim of this project is to assess whether the reporter cell line can be used for predicting toxicity of chemicals.

Project plan:

This project is a collaboration between Bioneer A/S and DTU Food, both located north of Copenhagen. We are collaborating already on the culture of iPSC and differentiation into cardiac cells. For this project, the gene editing will be done at Bioneer A/S, which has a large expertise with CRISPR/Cas9 and hiPSC, and recently generated other reporter cell lines. This is expected to take 6 – 7 months. After this, the reporter cell line will be transferred to DTU Food, and differentiated into cardiac cells with our recently developed protocol. Here, the cardiac differentiation and luciferase expression will be monitored using standard molecular methods, which will take 2 – 3 months. After this, luciferase expression will be monitored, and ultimately chemicals applied that impair cardiac differentiation. This will be done in the last months of the project. 

Qualifications:

The candidate is expected to have a solid background in molecular biology and cell biology. A high level of experience with standard techniques such as cloning, PCR and human cell culture is required. Knowledge in CRISPR/Cas9 is advantageous but not necessary. Experience with hiPSC culture is advantageous, but not a must. Most importantly, the candidate is expected to work independently and to be active in communication with all people involved. Very well written and spoken English is a must, Danish is not needed. We are looking for an open, active and skilled person, who is willing to learn a lot and conduct research on a high academic level.

We offer:

At DTU Food we are a big research group specialized in toxicology, ranging from in vivo to in vitro and computational approaches. We have a large variety of cell-based assays routinely running in the lab, and a high expertise in molecular analysis of the different cell types and assays. Bioneer A/S is a mid-sized Danish company with strong focus on research, offering training in state-of-the art facilities and supervision by experts within CRISPR/Cas9 and hiPSC culture.

All together, we offer the possibility to gain insights in both the private sector and academic research. This project is outstanding in the way that it gives the student the opportunity to learn numerous different techniques and assays, as well as cutting edge developments on the forefront of molecular and cell biology.

The student will be co-supervised by Karin Lauschke (DTU Food), Mikkel Rasmussen (Bioneer A/S), and the main supervisor will be Anne Marie Vinggaard (DTU Food).

Send your application letter and CV to Anne Marie Vinggaard, annv@food.dtu.dk




In collaboration with

Bioneer A/S

Requirements

The candidate is expected to have a solid background in molecular biology and cell biology. A high level of experience with standard techniques such as cloning, PCR and human cell culture is required.

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Contact

Company / Organization

DTU Fødevareinstituttet

Name

Anne Marie Vinggaard

Position

Professor

Mail

annv@food.dtu.dk

Supervisor info

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Technical University of Denmark

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