INDUCIBLE GENE TARGETING IN MICE EBOOK

Start studying Inducible Gene Targeting in Mice Use the Cre/lox System. Learn vocabulary, terms, and more with flashcards, games, and other study tools. Lessons on Conditional Gene Targeting in Mouse Adipose Tissue as well as a tamoxifen-inducible Cre driven by the aP2 gene promoter (iaP2), were bred to. In this paper, we provide a detailed protocol for tamoxifen-inducible gene deletion in neonatal mice, as well as for retina dissection.


INDUCIBLE GENE TARGETING IN MICE EBOOK

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INDUCIBLE GENE TARGETING IN MICE EBOOK

Inducible gene targeting in mice.

Advanced Search Abstract We have generated an optimized inducible recombination system for conditional gene targeting based on a Cre recombinase—steroid receptor fusion. Our findings provide a new concept allowing the generation of inducible mouse mutants within 6 months, as compared to 14 months using the current protocol.

Spatially regulated gene inactivation has proven to be powerful in dissecting the role of different cell types in a physiological process, as demonstrated for example by inducible gene targeting in mice concerning the role of insulin receptor signaling in glucose homeostasis 5.

The inducible gene targeting in mice of choice for precise gene function analyses in adult mice, however, is the temporal control of gene inactivation as it can prevent impaired embryonic development until the time of induction.

In particular, inducible gene targeting in all organs would be a useful tool if the major site of gene action is not precisely defined.

INDUCIBLE GENE TARGETING IN MICE EBOOK

Optimally, such a system should permit a tight control of inducible gene targeting in mice inactivation, efficient recombination in every single cell of the body upon induction, and the opportunity to produce sufficient numbers of inducible mouse mutants within a short time.

The major drawback of the current strategy is the long time frame required for the derivation of a conditional mouse mutant. While the first conditional mouse mutants can inducible gene targeting in mice expected within 14 months, additional time for breeding is usually needed to obtain sufficient numbers of mice for phenotypic analysis.

To date, only a few mouse strains permitting ubiquitously inducible gene targeting are available, all of which suffer from incomplete recombination upon induction or background activity in the absence of inducer 6 — Therefore, the side effects of the inducer may interfere with the phenotypic outcome of target gene inactivation.

In this paper we describe a novel approach for the rapid generation inducible gene targeting in mice inducible mouse mutants, which overcomes the drawbacks of the current technology.

In addition, we demonstrate that hybrid ES cells tolerate at least three consecutive gene targeting cycles including selection with the antibiotics G and hygromycin B without affecting the potency to complement tetraploid blastocysts.

Inducible Gene Targeting in Mice Using the Cre/loxSystem | Zenodo

Cell culture Culture and targeted mutagenesis of ES cells were carried out as previously described 15 with ES cell lines derived from both inbred and F 1 embryos. Generation of ect2floxES cells.

B6D2F1 mice for the generation of tetraploid blastocysts were obtained from Janvier. Production of ES mice by tetraploid embryo complementation The production of mice by inducible gene targeting in mice embryo complementation was essentially performed as described The CreERT2 coding region was inserted into the ubiquitously expressed Rosa26 locus through homologous recombination to obtain a configuration that allows the inducible inactivation of genes in all organs Fig.

We used the hybrid ES cell inducible gene targeting in mice V6. Deletion of the neo gene did not affect the level of CreERT2 expression as determined by western blot analysis Fig.

Rapid generation of inducible mouse mutants | Nucleic Acids Research | Oxford Academic

In contrast, the production of ES mice using inbred ES cells inducible gene targeting in mice inefficient 0. These results demonstrate that mice can be produced by tetraploid blastocyst complementation using cells that have undergone three consecutive rounds of transfection, including two drug selection cycles.

Ubiquitously inducible gene targeting in mice For the induction of CreER T2 in mice, we applied 1—5 mg tamoxifen orally for 5 days in the following experiments. It has been recently demonstrated that the oral application of tamoxifen is as efficient inducible gene targeting in mice i.

As a third substrate, we used a 2.

Inducible gene targeting in mice using the Cre/lox system.

A similar degree of recombination was achieved when a shorter 0. To investigate whether the low recombination efficiency in brain was due to low CreERT2 expression we performed western analysis using antibodies specific for the human estrogen receptor.

The 74 kDa band corresponding to the CreER T2 fusion protein was detectable in all organs including brain, with the highest inducible gene targeting in mice in thymus, small intestine and testis Fig.

INDUCIBLE GENE TARGETING IN MICE EBOOK

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