Talen technology used to produce a synovial sarcoma cell model

Details

Ressource 1Download: BIB_BCFD2D120384.P001.pdf (2419.57 [Ko])
State: Public
Version: After imprimatur
Secondary document(s)
Download: Mémoire no 2497 Annexes Mme El Chammah.pdf (935.55 [Ko])
State: Public
Version: author
Serval ID
serval:BIB_BCFD2D120384
Type
A Master's thesis.
Publication sub-type
Master (thesis) (master)
Collection
Publications
Institution
Title
Talen technology used to produce a synovial sarcoma cell model
Author(s)
EL CHAMMAH S.
Director(s)
STAMENKOVIC I.
Codirector(s)
FUSCO C.
Institution details
Université de Lausanne, Faculté de biologie et médecine
Publication state
Accepted
Issued date
2015
Language
english
Number of pages
21
Abstract
Synovial sarcoma (SS) is to this day a devastating malignancy, of which we have very little mechanistic understanding. Two elements characterize SS: the cell of origin, a multipotent myoblast precursor cell, and its signature chromosomal translocation t(X:18) that generates the SYT-SSX fusion gene and corresponding fusion protein. The aim of this project was to produce a TALEN, a tool that could be used to produce SS cells de novo by introducing the translocation into a selected cell line (C3H cells). This may give us insight into the translocation's impact on the cell's physiology early in transformation. The TALEN, composed of a nuclease (Folk I) and an amino acid repeat module array, responsible for the specificity of the nuclease's cut point, was generated using a basic cloning technique. The targeted break point in the murine genome was determined following several criteria including its resemblance to human SS break point and the limitation of off-target effect. Damaged DNA produced by the transfection of C3H cells with the TALEN triggered homologous recombination (HR) thereby introducing the LoxP site into the targeted sequence. The lox P sites were then recognized and recombined by CRE thus producing the translocation between chromosome X and 18. Efficiency and specificity of TALEN cleavage must now be demonstrated by PCR screening. HR's specificity and efficiency, must also be established using T7 Endonuclease I mutation detection assay.
Create date
31/08/2016 15:21
Last modification date
20/08/2019 16:31
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