Results 11 - 20 of 63

Targeting frequency for deletion vectors in embryonic stem cells

by H Zhang, P Hasty, A Bradley, Hongbing Zhang, Paul Hasty, Allan Bradleyl - GENE TRAPPING WITH DRUG-SELECTABLE MARKERS 9941 o n February 27, 2014 by PENN STATE UNIV , 1994
"... embryonic stem cells. Targeting frequency for deletion vectors in ..."
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embryonic stem cells. Targeting frequency for deletion vectors in

Potentiation of gene targeting in human cells by expression of Saccharomyces cerevisiae Rad52

by Cristina Di Primio, Alvaro Galli, Tiziana Cervelli , 2005
"... When exogenous DNA is stably introduced in mammalian cells, it is typically integrated in random positions, and only a minor fraction enters a pathway of homologous recombination (HR). The complex Rad51/Rad52 is a major player in the management of exogenous DNA in eukaryotic organisms and plays a cr ..."
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When exogenous DNA is stably introduced in mammalian cells, it is typically integrated in random positions, and only a minor fraction enters a pathway of homologous recombination (HR). The complex Rad51/Rad52 is a major player in the management of exogenous DNA in eukaryotic organisms and plays a critical role in the choice of repair system. In Saccharomyces cerevisiae, the pathway of choice is HR, mediated by Rad52 (ScRad52), which differs slightly from its human homologue. Here, we present an approach that utilizes ScRad52 to enhance HR in human cells containing a specific substrate for recombination. Clones of HeLa cells were produced expressing functional ScRad52. These cells showed enhanced resistance to DNA damaging treatments and revealed a different distribution of Rad51 foci (a marker of recombination complex formation). More significantly, ScRad52 expression resulted in an up to 37-fold increase in gene targeting by HR. In the same cells, random integration of exogenous DNA was significantly reduced, consistent with the view that HR and non-homologous end joining are alternative competing pathways. Expression of ScRad52 could offer a major improvement for experiments requiring gene targeting by HR, both in basic research and in gene therapy studies.
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Multi-kilobase homozygous targeted gene replacement in human induced pluripotent stem cells

by Susan M. Byrne, Luis Ortiz, Prashant Mali, John Aach, George M. Church , 2014
"... Sequence-specific nucleases such as TALEN and the CRISPR/Cas9 system have so far been used to disrupt, correct or insert transgenes at precise locations in mammalian genomes. We demonstrate efficient ‘knock-in ’ targeted replacement of multi-kilobase genes in human induced pluripotent stem cells (iP ..."
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Sequence-specific nucleases such as TALEN and the CRISPR/Cas9 system have so far been used to disrupt, correct or insert transgenes at precise locations in mammalian genomes. We demonstrate efficient ‘knock-in ’ targeted replacement of multi-kilobase genes in human induced pluripotent stem cells (iPSC). Using a model system replacing en-dogenous human genes with their mouse counter-part, we performed a comprehensive study of tar-geting vector design parameters for homologous re-combination. A 2.7 kilobase (kb) homozygous gene replacement was achieved in up to 11 % of iPSC with-out selection. The optimal homology arm length was around 2 kb, with homology length being especially critical on the arm not adjacent to the cut site. Homol-ogous sequence inside the cut sites was detrimental to targeting efficiency, consistent with a synthesis-dependent strand annealing (SDSA) mechanism. Us-ing two nuclease sites, we observed a high degree of gene excisions and inversions, which sometimes occurred more frequently than indel mutations. While homozygous deletions of 86 kb were achieved with up to 8 % frequency, deletion frequencies were not solely a function of nuclease activity and deletion size. Our results analyzing the optimal parameters for targeting vector design will inform future gene targeting efforts involving multi-kilobase gene seg-ments, particularly in human iPSC.
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Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster

by K J Keeler, T Dray, J E Penney, G B Gloor, Mol Cell Biol, Katherine J. Keeler, Tammy Dray, Janice E. Penney, Gregory, B. Gloor - Mol. Cell. Biol , 1996
"... Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster. ..."
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Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster.
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The effects of polymorphisms on human gene targeting

by David R. Deyle, Li B. Li, Gaoying Ren, David W. Russell - Nucleic Acids Research , 2013
"... DNA mismatches that occur between vector homology arms and chromosomal target sequences reduce gene targeting frequencies in several species; however, this has not been reported in human cells. Here we demonstrate that even a single mismatched base pair can significantly decrease human gene targetin ..."
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DNA mismatches that occur between vector homology arms and chromosomal target sequences reduce gene targeting frequencies in several species; however, this has not been reported in human cells. Here we demonstrate that even a single mismatched base pair can significantly decrease human gene targeting frequencies. In addition, we show that homology arm polymorph-isms can be used to direct allele-specific targeting or to improve unfavorable vector designs that intro-duce deletions.

Forecast Alert

by Keng Tee, Guihua Chen, Ling Lu, Wangxia Tong, Lin Gu, Chunhua Li, Teng Lu - IT Spending, Worldwide, 2008-2015, 4Q11 Update. 2012 http://www.gartner.com: http://www.gartner.com/it/content/1870000/1870018/january_10_it_spending_forecast_4q11_update_rgordon.p df?userId=57517080 , 2011
"... This article cites 42 articles, 15 of which can be accessed free ..."
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This article cites 42 articles, 15 of which can be accessed free

Precise BAC targeting of genetically polymorphic mouse ES

by Tahsin Stefan Barakat, Eveline Rentmeester, Frank Sleutels, J. Anton Grootegoed, Joost Gribnau , 2011
"... cells ..."
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Abstract not found
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In

by Linkage Groups, Hox A, Hox B, Hox C, Hox D, Which Are
"... vertebrates there is a matrix of 38 genes that collectively make up the Hox complex. These genes are distributed in four ..."
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vertebrates there is a matrix of 38 genes that collectively make up the Hox complex. These genes are distributed in four
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unknown title

by Valance Washington, Jun Cheng, Amalia Dutra, Evgenia Pak, Pentao Liu, Daniel W. Mcvicar, Pamela L. Schwartzberg , 2008
"... Advantages of q-PCR as a method of screening for gene targeting in mammalian cells using conventional and whole BAC-based constructs ..."
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Advantages of q-PCR as a method of screening for gene targeting in mammalian cells using conventional and whole BAC-based constructs
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Gene Targeting of a Plasmid-Borne Sequence to a Double- Strand DNA Break in Drosophila melanogaster

by Gregory B Gloor, Katherine J. Keeler, Tammy Dray, Janice E. Penney, Gregory, B. Gloor , 1995
"... Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster ..."
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Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster
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