scholarly journals Rescue of the TTF2 Knockdown Phenotype with an siRNA-Resistant Replacement Vector

Cell Cycle ◽  
2004 ◽  
Vol 3 (9) ◽  
pp. 1149-1151 ◽  
Author(s):  
Yan Jiang ◽  
David H. Price
Keyword(s):  
Replacement Vector

scholarly journals Target frequency and integration pattern for insertion and replacement vectors in embryonic stem cells

1991 ◽  
Vol 11 (9) ◽  
pp. 4509-4517
Author(s):  
P Hasty ◽  
J Rivera-Pérez ◽  
C Chang ◽  
A Bradley
Keyword(s):  
Homologous Recombination ◽  
Gene Targeting ◽  
Mammalian Cells ◽  
Germ Line ◽  
Es Cells ◽  
Embryonic Stem ◽  
Homologous Sequence ◽  
Reciprocal Recombination ◽  
Replacement Vector ◽  
Insertion Vector

Gene targeting has been used to direct mutations into specific chromosomal loci in murine embryonic stem (ES) cells. The altered locus can be studied in vivo with chimeras and, if the mutated cells contribute to the germ line, in their offspring. Although homologous recombination is the basis for the widely used gene targeting techniques, to date, the mechanism of homologous recombination between a vector and the chromosomal target in mammalian cells is essentially unknown. Here we look at the nature of gene targeting in ES cells by comparing an insertion vector with replacement vectors that target hprt. We found that the insertion vector targeted up to ninefold more frequently than a replacement vector with the same length of homologous sequence. We also observed that the majority of clones targeted with replacement vectors did not recombine as predicted. Analysis of the recombinant structures showed that the external heterologous sequences were often incorporated into the target locus. This observation can be explained by either single reciprocal recombination (vector insertion) of a recircularized vector or double reciprocal recombination/gene conversion (gene replacement) of a vector concatemer. Thus, single reciprocal recombination of an insertion vector occurs 92-fold more frequently than double reciprocal recombination of a replacement vector with crossover junctions on both the long and short arms.

scholarly journals Targeting the Root Cause of Mucopolysaccharidosis IIIA with a New scAAV9 Gene Replacement Vector

2020 ◽  
Vol 19 ◽  
pp. 474-485
Author(s):  
Tierra A. Bobo ◽  
Preston N. Samowitz ◽  
Michael I. Robinson ◽  
Haiyan Fu
Keyword(s):  
Gene Replacement ◽  
Root Cause ◽  
Replacement Vector

scholarly journals A caveat in mouse genetic engineering: ectopic gene targeting in ES cells by bidirectional extension of the homology arms of a gene replacement vector carrying human PARP-1

2008 ◽  
Vol 18 (2) ◽  
pp. 261-279 ◽  
Author(s):  
Aswin Mangerich ◽  
Harry Scherthan ◽  
Jörg Diefenbach ◽  
Ulrich Kloz ◽  
Franciscus van der Hoeven ◽  
...  
Keyword(s):  
Genetic Engineering ◽  
Gene Targeting ◽  
Es Cells ◽  
Gene Replacement ◽  
Replacement Vector ◽  
Parp 1 ◽  
Mouse Genetic

scholarly journals Integration of an insertion-type transferred DNA vector from Agrobacterium tumefaciens into the Saccharomyces cerevisiae genome by gap repair.

1996 ◽  
Vol 16 (10) ◽  
pp. 5924-5932 ◽  
Author(s):  
E Risseeuw ◽  
M E Franke-van Dijk ◽  
P J Hooykaas
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Agrobacterium Tumefaciens ◽  
Homologous Recombination ◽  
Model System ◽  
Transformation Frequency ◽  
Yeast Genome ◽  
Repair Mechanism ◽  
Replacement Vector ◽  
Dna Vector ◽  
Yeast Transformants

Recently, it was shown that Agrobacterium tumefaciens can transfer transferred DNA (T-DNA) to Saccharomyces cerevisiae and that this T-DNA, when used as a replacement vector, is integrated via homologous recombination into the yeast genome. To test whether T-DNA can be a suitable substrate for integration via the gap repair mechanism as well, a model system developed for detection of homologous recombination events in plants was transferred to S. cerevisiae. Analysis of the yeast transformants revealed that an insertion type T-DNA vector can indeed be integrated via gap repair. Interestingly, the transformation frequency and the type of recombination events turned out to depend strongly on the orientation of the insert between the borders in such an insertion type T-DNA vector.

A replacement vector used to introduce subtle mutations into mouse genes

Gene ◽  
1995 ◽  
Vol 166 (2) ◽  
pp. 197-204 ◽  
Author(s):  
Kyoji Horie ◽  
Shuichiro Maeda ◽  
Seiji Nishiguchi ◽  
Max E. Gottesman ◽  
Kazunori Shimada
Keyword(s):  
Replacement Vector

scholarly journals The Mechanism of Mammalian Gene Replacement Is Consistent with the Formation of Long Regions of Heteroduplex DNA Associated with Two Crossing-Over Events

2001 ◽  
Vol 21 (2) ◽  
pp. 501-510 ◽  
Author(s):  
Julang Li ◽  
Leah R. Read ◽  
Mark D. Baker
Keyword(s):  
Gene Replacement ◽  
Crossing Over ◽  
Replacement Reaction ◽  
Heteroduplex Dna ◽  
Mammalian Gene ◽  
Replacement Vector ◽  
Dna Strands ◽  
Vector Borne ◽  
Efficient Recovery ◽  
Vector Dna

ABSTRACT In this study, the mechanism of mammalian gene replacement was investigated. The system is based on detecting homologous recombination between transferred vector DNA and the haploid, chromosomal immunoglobulin μ-δ region in a murine hybridoma cell line. The backbone of the gene replacement vector (pCμCδpal) consists of pSV2neo sequences bounded on one side by homology to the μ gene constant (Cμ) region and on the other side by homology to the δ gene constant (Cδ) region. The Cμ and Cδ flanking arms of homology were marked by insertions of an identical 30-bp palindrome which frequently escapes mismatch repair when in heteroduplex DNA (hDNA). As a result, intermediates bearing unrepaired hDNA generate mixed (sectored) recombinants following DNA replication and cell division. To monitor the presence and position of sectored sites and, hence, hDNA formation during the recombination process, the palindrome contained a unique NotI site that replaced an endogenous restriction enzyme site at each marker position in the vector-borne Cμ and Cδ regions. Gene replacement was studied under conditions which permitted the efficient recovery of the product(s) of individual recombination events. Analysis of marker segregation patterns in independent recombinants revealed that extensive hDNA was formed within the Cμ and Cδ regions. In several recombinants, palindrome markers in the Cμ and Cδ regions resided on opposite DNA strands (trans configuration). These results are consistent with the mammalian gene replacement reaction involving two crossing-over events in homologous flanking DNA.

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