TITLE:
Targeted Mutagenesis of Beta-Lactoglobulin Gene in Caprine Fetal Fibroblasts by Context-Dependent Assembly Zinc-Finger Nucleases
AUTHORS:
Yuguo Yuan, Yong Cheng, Jinyu Wang, Qiuling Peng
KEYWORDS:
Context-Dependent Assembly, Zinc Finger Nucleases, Beta-Lactoglobulin, Gene Knockout, Goat
JOURNAL NAME:
Open Access Library Journal,
Vol.3 No.7,
July
26,
2016
ABSTRACT: Targeted mutagenesis by
zinc-finger nucleases (ZFNs) can be used to generate knock-out mammalian cell
lines with high efficiency. A number of different methods have been developed
for the design and assembly of gene-specific ZFNs, making them easily accessible
to researchers. In this study, we used ZFNs assembled through the CoDA
(context-dependent assembly) platforms to generate mutant caprine fetal fibroblasts
cells for the BLG gene. ZFN plasmid was introduced into the caprine fetal
fibroblasts cell by electroporation. ZFN-induced cleavage of the target
sequence was confirmed by the Surveyor nuclease assay analysis. Sequence
analysis revealed that ZFN-induced mutations such as base insertion,
deletion, or substitution were generated in the ZFN cleavage site of BLG. The
simplicity and efficacy of CoDA will enable broad application of ZFN
technology. This technique could be used with homologous arm, which may target
foreign genes into the BLG locus at higher efficiency.