Cre-mediated site-specific translocation between nonhomologous mouse chromosomes.

Deursen, Jan van; Fornerod, Maarten; Rees, Bastiaan van; Grosveld, Gerard

doi:10.1073/pnas.92.16.7376

Cited by 140 publications

(77 citation statements)

References 20 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…However, PCR analysis of a pool of 200 clones did not show unambiguous evidence for the desired inter-chromosomal rearrangement (data not shown). This is in agreement with the ®ndings of other studies, which have reported translocation frequencies of less than 1 in 1000 (van Deursen et al, 1995;Smith et al, 1995). It is unclear at present, why the intrachromosomal rearrangement occurs at a much higher frequency than the inter-chromosomal event.…”

Section: *Correspondence: T Boehmsupporting

confidence: 92%

“…A further signi®cant advantage of this strategy is that deletions can be generated in vivo. Previously, Cre-loxP recombination has been used to generate site-directed chromosomal translocations (Smith et al, 1995;van Deursen et al, 1995) and large deletions (Ramirez-Solis et al, 1995;Li et al, 1996). Our deletion strategy extends these ®ndings by incorporating the facility to recover intervening sequences in clonable form.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Predetermined chromosomal deletion encompassing the Nf-1 gene

et al. 1999

View full text Add to dashboard Cite

Complex chromosomal rearrangements (deletions, inversions, translocations) are a hallmark of human tumour cells. Yet, the generation of animal models for gross chromosomal abnormalities still presents a formidable challenge. Here, we describe a versatile procedure for chromosomal engineering that was used to generate an ES cell line with a megabase deletion encompassing the tumour suppressor gene neuro®bromatosis-1 (Nf-1) on mouse chromosome 11, which is often deleted in tumours of neural crest origin. Homologous recombination into sites¯anking Nf-1 was used to introduce arti®cial sequences (triple-helix, loxP, vector backbone) that can be employed for in vitro recovery of intervening sequences or the generation of in vivo deletions. This strategy may be developed into a scheme by which large chromosomal regions with precisely de®ned end points may be excised from mammalian cells and reintroduced after suitable in vitro modi®cation.

show abstract

Section: *Correspondence: T Boehmsupporting

confidence: 92%

mentioning

confidence: 99%

Predetermined chromosomal deletion encompassing the Nf-1 gene

et al. 1999

View full text Add to dashboard Cite

show abstract

“…De novo chromosomal translocations in mice using cre-loxP recombination An alternative approach to mimicing chromosomal translocations is the use of the Cre-loxP recombination system in ES cells to induce de novo chromosomal translocations (Smith et al, 1995;van Deursen et al, 1995). This has the advantage that tissue speci®c expression of the Cre recombinase would allow the translocation to occur in a tissue speci®c manner and, in addition, the de novo chromosomal translocation Figure 3 Lmo2 expression in vascularization of mouse embryos.…”

Section: An Mll-af9gene Fusion Knock-in Tumorigenesis Modelmentioning

confidence: 99%

Chromosomal translocation master genes, mouse models and experimental therapeutics

Rabbitts

2001

Oncogene

View full text Add to dashboard Cite

Molecular biologists have elucidated general principles about chromosomal translocations by cloning oncogenes or fusion genes at chromosomal translocation junctions. These genes invariably encode intracellular proteins and in acute cancers, often involve transcription and developmental regulators, which are master regulators of cell fate (e.g. LMO2 which is involved in acute leukaemia). Chromosomal translocations are usually associated with speci®c cell types. The reason for this close association is under investigation using mouse models. We are trying to emulate the cell-speci®c consequences of chromosomal translocations in mice using homologous recombination in embryonic stem cells to generate de novo chromosomal translocations or to mimic the consequence of these translocations. In addition, chromosomal translocation genes and their products are important targets for therapy. We have designed new therapeutic strategies which include antigen-speci®c recruitment of endogenous cellular pathways to a ect cellular viability and a novel structured form of antisense to ablate the function of fusion mRNAs. We will evaluate these procedures in the mouse models of chromosomal translocations and the long term aim is to perfect rapid procedures for characterizing patient-speci®c chromosomal translocations to tailor therapy to individual patients. Oncogene (2001) 20, 5763 ± 5777.

show abstract

“…The sequence-specific manipulation of DNA by Cre recombinase represents a powerful approach to studying gene function. Cre has been used to conditionally express, mutagenize, and replace genes (15)(16)(17)(18)(19)(20), as well as rearrange chromosomes (21)(22)(23)(24) in mice, resulting in major advances in the understanding of gene function. The requirement for preexisting genomic loxP sites complicates the use of Cre in studies of gene function and is an impediment to the use of Cre in applications such as gene therapy.…”

Section: Design Of Unnatural Loxp Sites and Cre Variant Librariesmentioning

confidence: 99%

Directed evolution of the site specificity of Cre recombinase

Santoro

Schultz²

2002

Proc. Natl. Acad. Sci. U.S.A.

157

109

View full text Add to dashboard Cite

Cre recombinase from bacteriophage P1 recognizes a 34-bp recombination site, loxP, with exquisite sequence specificity and catalyzes the site-specific insertion, excision, or rearrangement of DNA. To better understand the molecular basis of protein-DNA recognition and generate recombinases with altered specificities, we have developed a directed evolution strategy that can be used to identify recombinases that recognize variant loxP sites. To be selected, members of a library of Cre variants produced by targeted random mutagenesis must rapidly catalyze recombination, in vivo, between two variant loxP sites that are located on a reporter plasmid. Recombination results in an altered pattern of fluorescent protein expression that can be identified by flow cytometry. Fluorescence-activated cell sorting can be used either to screen positively for recombinase variants that recognize a novel loxP site, or negatively for variants that cannot recognize the wild-type loxP site. The use of positive screening alone resulted in a relaxation of recombination site specificity, whereas a combination of positive and negative screening resulted in a switching of specificity. One of the identified recombinases selectively recombines a novel recombination site and operates at a rate identical to that of wild-type Cre. Analysis of the sequences of the resulting Cre variants provides insight into the evolution of these altered specificities. This and other systems should contribute to our understanding of protein-DNA recognition and may eventually be used to evolve custom-tailored recombinases that can be used for gene study and inactivation.S ite-specific recombinases carry out a multitude of critical functions in nature ranging from gene rearrangement to genome segregation. Cre, a 38-kDa recombinase from bacteriophage P1 that resolves genome dimers into monomers (1, 2), is one of the simplest and best understood of known recombinases. Like other members of the integrase family, Cre catalyzes recombination between two identical double-stranded DNA sites of a particular sequence (1-3). The enzyme requires no accessory proteins or cofactors and functions efficiently under a wide variety of cellular and noncellular conditions. The recombination site recognized by Cre is a 34-bp doublestranded DNA sequence known as loxP (Fig. 1). The loxP site is palindromic with the exception of its eight innermost base pairs, which impart directionality to the site. Crystallographic analyses of Cre-DNA complexes (4-6) have helped to elucidate details of the catalytic mechanism of Cre-mediated site-specific recombination and identify protein-protein and protein-DNA interactions within the Cre-loxP catalytic complex. In a productive Cre-loxP complex, two loxP sites are aligned in an antiparallel orientation and are bound by four identical Cre subunits that join to form a ring in which each subunit contacts two adjacent subunits and one loxP half site.Cre can catalyze DNA integration, excision, or rearrangement, depending on the relative location and ori...

show abstract

Cre-mediated site-specific translocation between nonhomologous mouse chromosomes.

Cited by 140 publications

References 20 publications

Predetermined chromosomal deletion encompassing the Nf-1 gene

Predetermined chromosomal deletion encompassing the Nf-1 gene

Chromosomal translocation master genes, mouse models and experimental therapeutics

Directed evolution of the site specificity of Cre recombinase

Contact Info

Product

Resources

About