Target chromosomes of inducible deletion by a Cre/inverted loxP system in mouse embryonic stem cells

Tada, Masako; Matsumura, Hiroyuki; Kurose, Yuko; Nakatsuji, Norio; Tada, Takashi

doi:10.1007/s10577-009-9035-0

Cited by 10 publications

(7 citation statements)

References 15 publications

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“…In the second approach an active removal of the chromosome by exploiting cre-mediated rearrangements was proposed [ 19 ]. We tried to use this strategy, but the unexpected high frequency of the spontaneous loss of the endogenous engineered chromosome prevented us from implementing this approach, since normal substituted XY clones were easily obtained.…”

Section: Discussionmentioning

confidence: 99%

Chromosome transplantation as a novel approach for correcting complex genomic disorders

et al. 2015

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Genomic disorders resulting from large rearrangements of the genome remain an important unsolved issue in gene therapy. Chromosome transplantation, defined as the perfect replacement of an endogenous chromosome with a homologous one, has the potential of curing this kind of disorders. Here we report the first successful case of chromosome transplantation by replacement of an endogenous X chromosome carrying a mutation in the Hprt gene with a normal one in mouse embryonic stem cells (ESCs), correcting the genetic defect. The defect was also corrected by replacing the Y chromosome with an X chromosome. Chromosome transplanted clones maintained in vitro and in vivo features of stemness and contributed to chimera formation. Genome integrity was confirmed by cytogenetic and molecular genome analysis. The approach here proposed, with some modifications, might be used to cure various disorders due to other X chromosome aberrations in induced pluripotent stem (iPS) cells derived from affected patients.

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Section: Discussionmentioning

confidence: 99%

Chromosome transplantation as a novel approach for correcting complex genomic disorders

et al. 2015

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“…The recombination event appeared to be cytotoxic in the specific tissue analyzed (24). The same model has also been successfully utilized in chromosome deletion in embryonic stem cells (25)(26)(27). It was not known whether these detrimental effects apply to all adult tissue types and chromosomes.…”

Section: Discussionmentioning

confidence: 99%

Generation and Analysis of Partially Haploid Cells with Cre-mediated Chromosome Deletion in the Lymphoid System

Zhu

Kim

et al. 2010

Journal of Biological Chemistry

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The fast accumulation of mutant mouse strains in recent years has provided an invaluable resource for phenotype-based genetic screens. However, study of lymphoid phenotypes can be obscured or impractical if homozygous mutations cause early embryonic defects. To aid phenotype screening of germ line mutations in the lymphoid system, we developed a method to induce loss of heterozygosity (LOH) in developing lymphocytes through chromosome deletion. Chromosome deletion was triggered by Cre/loxP-mediated inverse sister chromatid recombination in the G 2 /M phase of the cell cycle, leading to the generation of daughter cells missing part of or the entire recombinant chromosome. We show that the resulting cells were viable and capable of additional rounds of cell division, thus providing raw materials for subsequent phenotypic assessment. We used the recombination system to induce LOH at the E2A locus in developing B cells. A significant loss of pro-B and pre-B cells was observed when the wild-type allele was removed by chromosome deletion from the E2A heterozygous mice, a result consistent with the required role for E2A in B cell development. We also demonstrated the effectiveness of Cre-mediated chromosome deletion in the LOH assay for HEB function in T cell development. Thus, the Cre-mediated chromosome deletion provides a new and effective method for genome-wide assessment of germ line mutations in the lymphoid system. Over 4000 protein-coding genes have been mutated and studied in the mouse by gene targeting, gene trapping, or other mutagenic methods in the past 2 decades (1, 2). This number is expected to increase dramatically following the recent calls for genome-scale coverage of germ line mutations by several international consortia (3). Studies of the existing germ line mutations have implied that approximately one-third of null mutations cause embryonic or neonatal lethality (4). Consequently, the effect of germ line mutations on postnatal life cannot be easily assessed for the genes playing important roles in both embryonic and postnatal life. Further analysis of tissue-specific gene function often requires a completely independent effort to establish a new allele for Cre/ lox-mediated conditional gene knock-out. Although the Cre/lox system can effectively address the embryonic problem and permit the study of tissue-specific gene function, the experimental system is not practical for large-scale genetic analysis. Thus far, there is no simple method available for systematic evaluation of lymphoid phenotypes of lethal mutations accumulated from a large-scale mutagenesis approach.Mitotic recombination provides one possible way to assess somatic phenotypes of pre-existing mutations. A recombination between homologous chromosomes during mitosis can lead to clonal segregation of heterologous alleles. The mosaics resulting from mitotic recombination allow for functional analysis of homozygous clones in an otherwise heterozygous background (5, 6). The feasibility of mosaic analysis in mice was demonstrated in the lo...

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“…However, recent studies using genetic engineering have revealed the possibility of performing aneuploidy therapy in cultured cells. Here, we summarize some of the main studies aimed at eliminating an entire chromosome using the Cre/loxP system [79][80][81][82][83][84][85], the TKneo transgene for positive and negative antibiotic selection [86], CRISPR/Cas9 system-mediated multiple cleavage [87,88], and zinc finger nuclease (ZFN)-mediated knock-in of the XIST gene to silence one copy of chromosome 21 [89] (cited in Table 2).…”

Section: Gene Targeting-mediated Chromosome Eliminationmentioning

confidence: 99%

Applications of Genome Editing Technology in Research on Chromosome Aneuploidy Disorders

Akutsu

Fujita

Tomioka

et al. 2020

Cells

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Chromosomal segregation errors in germ cells and early embryonic development underlie aneuploidies, which are numerical chromosomal abnormalities causing fetal absorption, developmental anomalies, and carcinogenesis. It has been considered that human aneuploidy disorders cannot be resolved by radical treatment. However, recent studies have demonstrated that aneuploidies can be rescued to a normal diploid state using genetic engineering in cultured cells. Here, we summarize a series of studies mainly applying genome editing to eliminate an extra copy of human chromosome 21, the cause of the most common constitutional aneuploidy disorder Down syndrome. We also present findings on induced pluripotent stem cell reprogramming, which has been shown to be one of the most promising technologies for converting aneuploidies into normal diploidy without the risk of genetic alterations such as genome editing-mediated off-target effects.

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Target chromosomes of inducible deletion by a Cre/inverted loxP system in mouse embryonic stem cells

Cited by 10 publications

References 15 publications

Chromosome transplantation as a novel approach for correcting complex genomic disorders

Chromosome transplantation as a novel approach for correcting complex genomic disorders

Generation and Analysis of Partially Haploid Cells with Cre-mediated Chromosome Deletion in the Lymphoid System

Applications of Genome Editing Technology in Research on Chromosome Aneuploidy Disorders

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