Gene targeting using zinc finger nucleases

2005 ◽  
Vol 23 (8) ◽  
pp. 967-973 ◽  
Author(s):  
Matthew H Porteus ◽  
Dana Carroll
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases

scholarly journals Comparing Zinc Finger Nucleases and Transcription Activator-Like Effector Nucleases for Gene Targeting in Drosophila

2013 ◽  
Vol 3 (10) ◽  
pp. 1717-1725 ◽  
Author(s):  
Kelly J. Beumer ◽  
Jonathan K. Trautman ◽  
Michelle Christian ◽  
Timothy J. Dahlem ◽  
Cathleen M. Lake ◽  
...  
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases ◽  
Transcription Activator

scholarly journals Efficient gene targeting in Drosophila by direct embryo injection with zinc-finger nucleases

2008 ◽  
Vol 105 (50) ◽  
pp. 19821-19826 ◽  
Author(s):  
K. J. Beumer ◽  
J. K. Trautman ◽  
A. Bozas ◽  
J.-L. Liu ◽  
J. Rutter ◽  
...  
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases ◽  
Efficient Gene

Zinc finger nucleases-mediated gene targeting in plant cells

2010 ◽  
Vol 150 ◽  
pp. 474-475
Author(s):  
A Tovkach ◽  
V Zeevi ◽  
D Weinthal ◽  
T Tzfira
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Plant Cells ◽  
Zinc Finger Nucleases

Gene Targeting in Drosophila and Caenorhabditis elegans With Zinc-Finger Nucleases

2008 ◽  
pp. 63-77 ◽  
Author(s):  
Dana Carroll ◽  
Kelly J. Beumer ◽  
J. Jason Morton ◽  
Ana Bozas ◽  
Jonathan K. Trautman
Keyword(s):  
Caenorhabditis Elegans ◽  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases

236 GENE TARGETING USING ZINC FINGER NUCLEASES (ZFN) IN THE PORCINE FIBROBLAST

2012 ◽  
Vol 24 (1) ◽  
pp. 230
Author(s):  
S. Kim ◽  
J. W. Kim ◽  
S. M. Lee ◽  
J. H. Kim ◽  
M. J. Kang
Keyword(s):  
Homologous Recombination ◽  
Gene Targeting ◽  
Zinc Finger ◽  
Marker Gene ◽  
Somatic Cells ◽  
Embryonic Stem ◽  
Domestic Animals ◽  
Zinc Finger Nucleases ◽  
Exogenous Dna ◽  
Targeting Vector

Gene targeting is a genetic technique that utilises homologous recombination between an engineered exogenous DNA fragment and the endogenous genome of an animal. In domestic animals, gene targeting has provided an important tool for producing knockout pigs for the α1,3-galactosyltransferase gene (GGTA1) to use in xenotransplantation. The frequency of homologous recombination is a critical parameter for the success of gene targeting. The efficiency of homologous recombination in somatic cells is lower than that in mouse embryonic stem cells. The application of gene targeting to somatic cells has been limited by its low efficiency. Recently, knockout rat and mouse were generated by introducing nonhomologus end joining (NHE)-mediated deletion or insertion at the target site using zinc-finger nucleases (ZFN). Therefore, the development of effective knockout and knock-in techniques in domestic animals is very important in biomedical research. In this study, we investigated homologous recombination events at the cytidine monophospho-N-acetylneuraminic acid hydroxylase (CMAH) gene locus using ZFN in porcine primary fibroblast. The CMAH-targeted ZFN plasmid and mRNA were purchased from Sigma-Aldrich (St Louis, MO, USA). Porcine ear fibroblasts cells were obtained from a 10-day-old male Chicago miniature pigs. The fibroblasts were cultured in DMEM containing 15% fetal bovine serum, 1 × nonessential amino acids, 1 × sodium pyruvate, 10–4 M β-mercaptoethanol, 100 unit mL–1 penicillin and 100 μg mL–1 streptomycin. The cells were trypsinized and resuspended at a concentration of 1.25 × 107 cells mL–1 in F10 nutrient mixture. Four hundred microliters of the cell suspension was electroporated in a 4-mm cuvette with 4 pulses of 1 ms duration using 400V capacitive discharges using the CMAH neo targeting vector and ZFN plasmid or RNA. The CMAH neo targeting vector consists of the neomycin resistance gene (neo) as a positive selectable marker gene, 789-bp 5′ arm and 763-bp 3′ arm from exon 8 of CMAH gene. After selection of G-418, PCR analysis was performed using 64 colonies transfected with ZFN plasmid and 48 colonies transfected with ZFN RNA. As a result, 19 positive colonies were identified in colonies transfected with ZFN plasmid and 15 colonies were identified in colonies transfected with ZFN RNA. The targeting efficiency was 29.7 and 31.6% in the colonies transfected with ZFN plasmid and ZFN RNA, respectively. To our knowledge, this study provides the first evidence that the efficiency of gene targeting using ZFN was higher than that of conventional gene targeting in the porcine fibroblast. These cell lines may be used in production of CMAH knockouts for xenotransplantation.

scholarly journals Gene targeting to the ROSA26 locus directed by engineered zinc finger nucleases

2011 ◽  
Vol 40 (8) ◽  
pp. 3741-3752 ◽  
Author(s):  
Pablo Perez-Pinera ◽  
David G. Ousterout ◽  
Matthew T. Brown ◽  
Charles A. Gersbach
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases ◽  
Rosa26 Locus

scholarly journals Efficient Gene Targeting in Drosophila With Zinc-Finger Nucleases

Genetics ◽  
2006 ◽  
Vol 172 (4) ◽  
pp. 2391-2403 ◽  
Author(s):  
Kelly Beumer ◽  
Gargi Bhattacharyya ◽  
Marina Bibikova ◽  
Jonathan K. Trautman ◽  
Dana Carroll
Keyword(s):  
Gene Targeting ◽  
Zinc Finger ◽  
Zinc Finger Nucleases ◽  
Efficient Gene
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