Validating the involvement of AGFG1 in X chromosome inactivation through CRISPR/Cas9 genome editing

• Main Author: Zheng, Zihao
• Other Authors: Zhang Li-Feng
• Format: Final Year Project
• Language: English
• Published: 2018
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/74150
• Institution: Nanyang Technological University

X chromosome inactivation (XCI) is a phenomenon that occurs in mammals, whereby X chromosome is silenced in female to balance the X-linked gene dosage. Although XCI and the role of non-coding RNA Xist in XCI have been intensively studied, the exact mechanism and modulators of XCI are largely unclear and awaits to be elucidated. This study aims to validate the role of AGFG1, a gene involved in nucleocytoplasmic transport, in XCI. Through inducing double stranded breaks and frameshift mutation via CRISPR/Cas9 system on AV3.1.12, a transgenic male mouse embryonic stem cell line carrying inducible Xist, and 3F1, a female mouse embryonic stem cell line, the knock-out cell lines were successfully established. Alteration in viability of AGFG1 knock-out cell lines under expression of ectopic Xist and during the differentiation as compared to the control were observed. However, further experiments need to be conducted to confirm the involvement of AGFG1 in silencing of the X chromosome.