Production of Transgenic Rats Using Pregnant and Pseudopregnant Rats Prepared at a Breeding Farm

Kajiwara, Noriko; Sugiyama, Fumihiro; Goto, Yasumasa; Sugiyama, Yuko; Fukamizu, Akiyoshi; Uehara, Satoshi; Sugimura, Keiichi; Murakami, Katsuji; Hokao, Ryoji; Akahori, Fumiaki

doi:10.1538/expanim1978.42.3_463

Cited by 5 publications

(5 citation statements)

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“…However, the high magnitude g force on the pronuclear zygotes themselves was reported not to be detrimental to their subsequent developmental potential in rabbits [11], pigs [39] and cattle [38]. The reduction in the developmental potential of the DNA-injected zygotes is known in mice [2,3,19,22,34], rats [4,5,13,15,17,20,23,32], rabbits [8,21,30,31,39] and pigs [8,24,27,28,36] to be from one-third to one-half of nontreated zygotes. In the present study, the maximum number of zygotes transferred into each recipient was the same in the four examined species and most of the recipient animals (13 of 13, 100% in mice; 15 of 15, 100% in rats; 8/11, 73% in rabbits; 7 of 9, 78% in pigs) became pregnant by the surgical transfer of the DNAinjected zygotes.…”

Section: Discussionmentioning

confidence: 99%

“…In the production of transgenic mice it was reported that 0.5 to 10.0% of the DNA-microinjected zygotes developed into offspring carrying exogenous DNA [2,3,19,22,34]. A range of 0.4 to 4.8% of the DNAmicroinjected rat zygotes were reported to develop into transgenic rats [4,13,15,17,20,32], and in rabbits and pigs, 0.3 to 2.6% [1,18,21,30,31] and 0.3 to 1.4% [8,[27][28][29]36] of the DNA-microinjected zygotes developed into the transgenic animals, respectively. However, concluding that the efficiency of producing transgenic animals is dependent on the body size of the animals based on these reports is problematic, because a great variety of exogenous DNA constructs and technical protocols for DNA microinjection were employed in each laboratory.…”

Section: Introductionmentioning

confidence: 99%

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A Comparative Study on the Integration of Exogenous DNA into Mouse, Rat, Rabbit, and Pig Genomes.

et al. 2001

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Transgenic mammals, from small laboratory rodents to domestic animals, have been successfully produced to date, but their production efficiency within or across species has been variable. This is probably due to the differences in the type of injected DNA and/ or technical procedures employed in each laboratory, as well as the reproductive characteristics of the species. Here we report the direct comparison of the efficiencies of producing transgenic mice, rats, rabbits and pigs by one technician using a fusion gene composed of the bovine αS 1 -casein promoter and human growth hormone (hGH) gene. Before the fusion gene was injected into the zygotes, high magnitude centrifugation to visualize the pronuclei was necessary for all of the pig zygotes and one-third of the rabbit zygotes, but not for mouse and rat zygotes. Post-injection survival of the mouse zygotes (67.1%) was lower than those of the rat, rabbit and pig zygotes (89.6 to 100%). The volume change of the pronucleus following DNA injection was the lowest in mice (50% increase), moderate in rabbits (148% increase), and the most prominent in rats (238% increase). The data from only 1 pig zygote indicated a 22% increase in the pronucleus volume by DNA injection. The PCR analyses of the tail DNA of new born offspring indicated that 0.8% (4/ 493), 4.8% (22/463), 0.8% (3/367) and 0.9% (2/221) of the injected eggs in mice, rats, rabbits and pigs, respectively, developed into transgenic offspring. Some of the founder animals in all four species expressed the transgene in the mammary gland which was confirmed in hGH mRNA by RT-PCR and/or hGH peptide in Witch's milk with ELISA. These results suggest that the maximum volume of DNA solution injectable into the pronucleus is a possible factor explaining the species differences in the production of transgenic animals.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A Comparative Study on the Integration of Exogenous DNA into Mouse, Rat, Rabbit, and Pig Genomes.

et al. 2001

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“…Injection of immature SD female rats with low doses of PMSG (10 IU) yields 16 +_ 5 viable eggs per animal, whereas higher doses of this gonadotropin produce a variable ovulatory response and abnormal embryo development (Walton et al, 1983;Young et al, 1987;Armstrong and Opavski, 1988), probably due to the relatively high LH-like activity of PMSG. Although less efficient in rats than in mice, this protocol has nevertheless been used successfully to produce transgenic rats (Hochi et al, 1990;Kajiwara et al, 1993) (Table 1). …”

Section: Superovulationmentioning

confidence: 99%

“…An effective alternative to PMSG for inducing superovulation in rats and producing large numbers of normally developed ova has been described by Armstrong and Opavski (1988) and has been used by several groups to obtain transgenic rats ( 0.5 Hammer et al, 1990Mullins et al, 1990Hochi et al, 1990Hochi et al, 1990Ganten et al, 1992Ganten et al, 1992Hochi et al, 1992Swanson et al, 1992Southard et al, 1992Matsumoto et al, 1993Kajiwara et al, 1993Dycaieo et al, 1994Charreau et al, (1996 also be sought on paper placed under the grid floor of the cages. Alternatively, mating can be confirmed by examining vaginal smears for sperm (Szabo et al, 1969).…”

Section: Superovulationmentioning

confidence: 99%

Transgenesis in rats: Technical aspects and models

Charreau

Tesson

Soulillou

et al. 1996

Transgenic Research

136

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The production of transgenic rats by DNA-microinjection into fertilized ova has now become an established procedure, although fewer than 20 lines have been described during the last 5 years. Overall, transgenic rats remain more difficult to produce than transgenic mice, but satisfactory yields have been obtained by several laboratories. A review of the methods used to generate transgenic rats shows considerable variation between different laboratories, particularly in choice of strain, superovulation protocols and the use of embryo culture before reimplantation. In some instances, the production of transgenic rats has provided data that are new and relevant, compared to data obtained in mice bearing the same transgene. Models have been developed for human diseases such as hypertension and autoimmunity, and applications have been found in the study of carcinogenesis and in pharmacological research. Transgenic rat technology also opens up interesting perspectives for transplantation research, in which microsurgery is an essential procedure. Intensive research is in progress in several laboratories to produce rat embryonic stem (ES) cell lines, but existing lines have not participated in germ line formation a prerequisite for their use in gene knock out experiments.

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“…Generally, it takes approximately 2 to 3 weeks until post-pubertal rats exhibit stable estrus cycles with a defined 4-day period. Fully matured rats, superovulated by injections of pregnant mare serum gonadotrophin (PMSG) at the diestrus-stage and of human chorionic gonadotrophin (hCG) at the proestrus-stage, have been used as donors for the production of transgenic rats [9,10]. In the preparation of embryo recipients, matured rats at the proestrus-stage must be mated with vasectomized male rats to induce pseudopregnancy.…”

Section: Introductionmentioning

confidence: 99%

Production of Transgenic Rats Using Young Sprague-Dawley Females Treated with PMSG and hCG.

et al. 2001

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Abstract:The aim of this study was to examine the effects of gonadotrophin treatments on estrus synchronization and superovulation in young Sprague-Dawley (SD) rats that had not yet exhibited defined estrus cycles (5 to 7 weeks old), and to produce transgenic rats using these females as embryo donors and recipients. In Experiment 1, female rats were injected with PMSG and hCG (12.5, 25, 50 and 100 IU/kg each) and were mated with stud males. The reproductive performance of young rats were highest when PMSG and hCG at doses of 25 IU/kg each were injected (delivery rate 87.5%, nursing rate 92.9%). In Experiment 2, female rats were injected with PMSG and hCG (100, 150 and 300 IU/kg each) to induce superovulation. More eggs were recovered from the rats injected with PMSG and hCG at 150 and 300 IU/kg than from those treated with 100 IU/kg (33.4 and 41.3 vs. 13.3 eggs per female, respectively; p<0.05). In Experiment 3, pronuclear-stage zygotes from 150 IU/kg PMSG/hCG-treated rats were used for microinjection of the fusion gene of bovine αS1-casein gene promoter and human growth hormone gene (2.8 kb), and the microinjected zygotes were transferred into the oviduct ampullae of the 25 IU/kg PMSG/hCG-treated rats. Seventeen transgenic rats were obtained from the 334 DNA-injected zygotes (5.1%). These results indicate that recipients and embryo donors for the production of transgenic rats can be prepared by the appropriate PMSG and hCG treatments of young SD rats, regardless of their estrus stages.

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Production of Transgenic Rats Using Pregnant and Pseudopregnant Rats Prepared at a Breeding Farm

Cited by 5 publications

References 5 publications

A Comparative Study on the Integration of Exogenous DNA into Mouse, Rat, Rabbit, and Pig Genomes.

A Comparative Study on the Integration of Exogenous DNA into Mouse, Rat, Rabbit, and Pig Genomes.

Transgenesis in rats: Technical aspects and models

Production of Transgenic Rats Using Young Sprague-Dawley Females Treated with PMSG and hCG.

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