Maintenance of Transgenes in Rats: The Contributions of Embryo Cryopreservation and Intracytoplasmic Sperm/Spermatid Injection.

Hochi, Shinichi; Hirabayashi, Masumi

doi:10.1262/jrd.48.205

Cited by 9 publications

(7 citation statements)

References 45 publications

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“…Pregnancy rates of foster mothers after embryo transfer of intact or microinjected zygotes were not different. The proportion of microinjected zygotes resulting in live offspring was ranging from 16.7 to 38.7% and was thus comparable to corresponding literature data (Charreau et al, 1996;Hirabayashi et al, 2001a;Hochi & Hirabayashi, 2002;Kato et al, 2004b). Surprisingly, full-term development of microinjected zygotes of the LEW strain transferred to foster mothers was higher compared to the other strains.…”

Section: Overall Efficiency Of Transgenesissupporting

confidence: 86%

“…The most important one is the skill of the scientist performing microinjections. It is known that pronuclear microinjection in rats is relative difficult compared to the mouse and there seems to be a 'knack' in microinjecting exogenous DNA solution into the pronucleus of rat zygotes (Hochi & Hirabayashi, 2002). In addition, the purification techniques of DNA for injection, the DNA concentration, the method used for microinjection and other factors are also very important for the outcome of transgenic technology (Wall, 2001).…”

Section: Overall Efficiency Of Transgenesismentioning

confidence: 99%

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Strain Differences in Superovulatory Response, Embryo Development and Efficiency of Transgenic Rat Production

2005

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The differences between rat strains in superovulation response, in vitro and in vivo development of preimplantation embryos and overall transgenic efficiency was studied. The protocols for induction of superovulation using single injections of pregnant mare's serum gonadotropin (PMSG) or minipumps with follicle stimulating hormone (FSH) were compared in Lewis (LEW), Wistar-Kyoto (WKY), and stroke-prone spontaneously hypertensive rats (SHRSP) or Sprague-Dawley (SD) and Wistar rats as representative inbred or outbred strains, respectively. The percentage of mated animals with positive superovulatory response was similar in all strains (60.0-100%). The mean number of ova per donor was not dependent on the kind of hormonal treatment used within each rat strain. In general, females from outbred SD and Wistar rats were more responsive to hormonal treatments than animals from inbred rat strains. In addition, SD female rats produced a significantly higher number of embryos per female in response to PMSG-treatment compared to all other strains. Between the inbred strains, SHRSP was the most effective for superovulation. In vitro development of intact zygotes to the blastocyst stage was not different between SD, Wistar and SHRSP rats. In contrast, in vitro development of WKY zygotes was significantly less efficient than in other strains. However, 2-cell stage embryos in vivo produced from SD, SD x Wistar and WKY animals showed no difference in competence to develop to blastocyst stage in vitro. The proportion of offspring developing after oviduct transfer of intact zygotes was similar in all strains (44.0-56.4%) with the exception of WKY rats (35.9%). We also compared the survival rate after injection, ability of manipulated zygotes to develop to term and overall transgenic efficiency in various rat strains. SD and SHRSP zygotes survived after microinjection better than the WKY and Lewis zygotes. No differences were found in the efficiency of transgene integration per newborn in different strains ranging from 5.7 to 16.7%. The results of this study demonstrate that different rat strains have varying responses to superovulation, sensitivity to microinjection, capability to develop in vitro until blastocyst stage or in vivo to term after transfer to foster mothers. Despite these differences all studied strains can be used for efficient transgenic rat production.

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Section: Overall Efficiency Of Transgenesissupporting

confidence: 86%

Section: Overall Efficiency Of Transgenesismentioning

confidence: 99%

Strain Differences in Superovulatory Response, Embryo Development and Efficiency of Transgenic Rat Production

2005

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“…Moreover, the low rate of full-term development after ROSI [61] may be explained either by the abnormal pattern of paternal genome demethylation [36] or by the abnormal localization of methylated chromatin in the nucleoplasm of male pronucleus rather than the failure to completely demethylate the paternal genome [38]. On the other hand, in vitro production of rat zygotes is not efficient as in mouse, despite of the successful birth of ICSI-or ROSI-derived rat offspring [62]. Paternal genome active demethylation in IVF-and ICSI-derived rat zygotes occurred at a similar timing (6 to 10 h after fertilization), but to lesser extent compared to the in vivo-derived counterparts.…”

Section: Effect Of In Vitro Embryo Production On Active Demethylationmentioning

confidence: 99%

Active Demethylation of Paternal Genome in Mammalian Zygotes

Abdalla

Yoshizawa

Hochi

2009

Journal of Reproduction and Development

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Abstract. Epigenetic reprogramming in early preimplantation embryos, that refers to erasing and remodeling epigenetic marks such as DNA methylation, is essential for differentiation and development. In many species, paternal genome is subjected to genome-wide active demethylation before the DNA replication commences, while maternal genome maintains its methylation status until being demethylated passively during the subsequent cleavage divisions. The purpose of this manuscript was to review the available knowledge about the paternal genome active demethylation process concerning the possible mechanisms, species variation and the factors affecting the active demethylation dynamics such as in vitro protocols for production of pronuclear-stage zygotes. Better understanding the mechanisms by which the epigenetic reprogramming is occurred may contribute to clarify the biological significance of this process. Key words: Active demethylation, Epigenetics, Paternal genome, Pronuclear zygotes (J. Reprod. Dev. 55: [356][357][358][359][360] 2009) ene expression is regulated by both genetic and epigenetic mechanisms. Epigenetics includes modifications of DNA itself (methylation of cytosine in the CpG dinucleotide) and/or the associated protein (phosphorylation, acetylation and methylation of histone) [1]. These modifications can regulate the gene expression without changing the DNA consequences [2]. Each cell type in our body has its own epigenetic signature which reflects different gene expression, and subsequently results in different structure and function among genetically homogenous cells. DNA methylation is one of the most-studied epigenetic mechanisms, and it is recognized as a chief contributor to the stability of gene expression state [3]. DNA Methylation: Nature, Function and ReprogrammingDNA methylation is a process that includes transferring a methyl group from S-adenosylmethionine to C5 positions of the cytosine residues in the CpG dinucleotides by different categories of methyltransferase enzymes (Dnmt1, Dnmt1o, Dnmt2, Dnmt3a, Dnmt3b and Dnmt3L) [4,5]. The CpG dinucleotides are mainly present in a cluster called CpG islands, which are associated with genes, and mostly located in promoters and first exons [6]. These islands are defined as the initiation sites for both transcription and DNA replication, and may represent genomic footprints for replication initiation [7]. Methylation of the CpG dinucleotides in the gene promoter may repress gene expression by interfering with the access of the DNA binding proteins and subsequently blocking the transcription or by binding to the transcriptional repressor MeCP2 [4,8,9]. In spite of great correlation between gene methylation status and gene expression, it is not yet clear whether DNA methylation is the cause of or a subsequence to the transcriptional repression [10]. In an another view for the correlation between gene methylation status and its expression, methylation can increase or decrease the level of gene transcription depending on whether the methylation in...

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“…Isotani et al [33] recently reported that rat ES cells injected into nu/nu mouse blastocysts could contribute to form not only thymus but also sperm-like germ cells. Although the minimal essential techniques for microinsemination, such as intracytoplasmic sperm injection and round spermatid injection, are almost available in the rat [34], the functional normality of the rat germ celllike cells observed in such xenogenic chimeras has not yet been confirmed. Nevertheless, the interspecies blastocyst complementation system using mutant mice or experimentally nitch-induced mice and pluripotent stem cells of the rats would provide an appropriate model for generation of human germ cells in the body of non-human species.…”

Section: Organ Regeneration By Blastocyst Complementationmentioning

confidence: 99%

Embryonic stem (ES) cells and induced pluripotent stem (iPS) cells in rats

Hirabayashi

Hochi

2011

Reprod Medicine & Biology

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Over the past 25 years, the reverse genetic approach including precise and conditional replacement or loss of gene function at a specific locus was considered possible only in mice due to the absence of embryonic stem (ES) or induced pluripotent stem (iPS) cell lines in other species. Recently, however, stem cell technology in rats has become available for biomedical research. In this paper we overview the recent progress of rat ES and iPS cell technology. Starting from the establishment of rat ES cells, the use of ES cells for foreign gene transfer and endogenous gene knock-out is discussed, followed by the successful establishment of rat iPS cells and the generation of an iPS cell-derived organ via interspecific blastocyst complementation. Finally, the possible contribution of rat stem cell technology to reproductive medicine is described.

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Maintenance of Transgenes in Rats: The Contributions of Embryo Cryopreservation and Intracytoplasmic Sperm/Spermatid Injection.

Cited by 9 publications

References 45 publications

Strain Differences in Superovulatory Response, Embryo Development and Efficiency of Transgenic Rat Production

Strain Differences in Superovulatory Response, Embryo Development and Efficiency of Transgenic Rat Production

Active Demethylation of Paternal Genome in Mammalian Zygotes

Embryonic stem (ES) cells and induced pluripotent stem (iPS) cells in rats

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