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A Novel Method for Production of Androgenic Embryos Derived from Round Spermatids Injection in Mice

机译：小鼠圆形精子注射产生雄激素胚的新方法

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摘要

Mouse parthenogenetic/gynogenetic and androgenic embryos, which contain two sets of maternal and paternal haploid genomes, respectively, provide important materials for studies on the genomic imprinting. The present study was undertaken to observe the developmental potential of androgenetic embryos produced by ROSI technique. MII oocytes were collected from BDF1 or 129 females. Spermatogenic cells were isolated from BDF1, 129, or JF1 males. Two round spermatid nuclei were injected into TII oocyte, and the female nucleus was removed together with the second polar body. Survived embryos were cultured and those developing to the 2-cell stage were transferred into day 0.5 pseudopregnant ICR females. Dissections were carried out at day 9.5, and methylation of imprinted genes: H19, Igf2r, and IG-DMR were analyzed in recovered fetuses. When BDF1 and 129 oocytes were used, cleavage rates were 61.7% (930/1508) and 50.2% (196/360), respectively. The in vivo survival rates of embryos from BDF1 and 129 spermatids were 7.6% (7/91) and 7.3% (3/41) for BDF1 oocytes and 6.6% (16/243) and 3.8% (5/130) for 129 oocytes, respectively. However, with JF1 spermatids, the rate was significantly(P < 0.05) lower [0.2% (1/461), with BDF1 oocytes] as compared with those from BDF1 or 129. DNA methylation analysis showed that these conceptuses were derived solely from the male genome. Although technical improvements are still needed, this study indicated that androgenesis using round spermatids may be a useful tool for production of androgenic embryos with genetic variations.
机译：小鼠孤雌/生殖和雄激素胚胎分别包含两组母本和父本单倍体基因组，为研究基因组印迹提供了重要的材料。进行本研究以观察通过ROSI技术产生的雄激素胚胎的发育潜力。从BDF1或129位雌性中收集MII卵母细胞。从BDF1、129或JF1雄性分离出生精细胞。将两个圆形的精子细胞核注入TII卵母细胞，并将雌性核与第二个极体一起切除。培养存活的胚胎，并将发育至2-细胞阶段的胚胎转移至0.5天假孕ICR雌性中。在第9.5天进行解剖，并在回收的胎儿中分析印迹基因H19，Igf2r和IG-DMR的甲基化。当使用BDF1和129卵母细胞时，切割率分别为61.7％（930/1508）和50.2％（196/360）。 BDF1和129个精子细胞的胚胎的体内存活率对于BDF1卵母细胞为7.6％（7/91）和7.3％（3/41），对于129个卵母细胞为6.6％（16/243）和3.8％（5/130） ， 分别。然而，与来自BDF1或129的精子相比，使用JF1精子的比率显着降低（P <0.05）[0.2％（1/461），对于BDF1卵母细胞。] DNA甲基化分析表明，这些概念仅来自于男性基因组。尽管仍需要技术上的改进，但这项研究表明，使用圆形精子的雄激素生成可能是生产具有遗传变异的雄激素胚胎的有用工具。

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