Generation of induced pluripotent stem cells from a newly identified type of laminopathy - 'proof of principle'.

• Main Author: Chua, Bernice Hong Min.
• Other Authors: School of Biological Sciences
• Format: Final Year Project
• Language: English
• Published: 2011
• Subjects: DRNTU::Science::Biological sciences::Genetics
• Online Access: http://hdl.handle.net/10356/45136
• Institution: Nanyang Technological University

Laminopathies constitute a family of disease caused by mutations in the lamin proteins which includes lamin A/C and lamin B1/2. Recently, a new type of laminopathy characterised by inherited heart disease and limb abnormalities was identified. Patients with this disease are at risk of sudden cardiac arrest. In this project, our work focused on generating a disease model of this disease by inducing Induced Pluripotent Stem Cells (IPSCs) from the primary human fibroblasts of these patients, using two in-house designed doxycycline (dox)-inducible lentiviruses constructs: one with the four Yamanaka factors (Oct-3/4, Sox-2, C-myc and Klf-4) and the other with five reprogramming factors (Oct-3/4, Sox-2, C-myc, Klf-4 and Nanog). IPSCs were formed from IMR90 and aged-matched control fibroblasts with these two viruses constructs thereby demonstrating the efficiency of our virus system. Results from immuno-staining and alkaline phosphatase of the IMR90 IPSCs confirmed the pluripotency of the generated IPSCs. However, we did not manage to obtain IPSCs from the laminopathy patient‟s fibroblasts despite 2 rounds of virus infection due to their progressive cell death during puromycin selection.