School of Mathematics, Physics and Biological Engineering, Inner Mongolia University of Science & Technology, Baotou, Inner Mongolia, China.
In Vitro Cell Dev Biol Anim. 2011 Feb;47(2):95-103. doi: 10.1007/s11626-010-9363-6. Epub 2010 Nov 17.
The technique of interspecies somatic cell nuclear transfer, in which interspecies cloned embryos can be reconstructed by using domestic animal oocytes as nuclear recipients and endangered animal or human somatic cells as nuclear donors, can afford more opportunities in endangered animal rescue and human tissue transplantation, but the application of this technique is limited by extremely low efficiency which may be attributed to donor nucleus not fully reprogrammed by xenogenic cytoplasm. In this study, goat fetal fibroblasts (GFFs) were used as nuclear donors, in vitro-matured sheep oocytes were used as nuclear recipients, and a two-stage nuclear transfer procedure was performed to improve the developmental ability of goat-sheep interspecies clone embryos. In the first stage nuclear transfer (FSNT), GFFs were injected into the ooplasm of enucleated sheep metaphase-II oocytes, then non-activated reconstructed embryos were cultured in vitro, so that the donor nucleus could be exposed to the ooplasm for a period of time. Subsequently, in the second stage nuclear transfer, FSNT-derived non-activated reconstructed embryo was centrifuged, and the donor nucleus was then transferred into another freshly enucleated sheep oocyte. Compared with the one-stage nuclear transfer, two-stage nuclear transfer could significantly enhance the blastocyst rate of goat-sheep interspecies clone embryos, and this result indicated that longtime exposure to xenogenic ooplasm benefits the donor nucleus to be reprogrammed. The two-stage nuclear transfer procedure has two advantages, one is that the donor nucleus can be exposed to the ooplasm for a long time, the other is that the problem of oocyte aging can be solved.
种间体细胞核移植技术，即将异种克隆胚胎通过用家畜卵母细胞作为核受体和濒危动物或人类体细胞作为核供体来构建，为濒危动物拯救和人类组织移植提供了更多机会，但该技术的应用受到极低效率的限制，这可能归因于供体核未被异种细胞质完全重编程。在这项研究中，山羊胎儿成纤维细胞（GFF）被用作核供体，体外成熟的绵羊卵母细胞被用作核受体，并采用两阶段核移植程序来提高山羊-绵羊种间克隆胚胎的发育能力。在第一阶段核移植（FSNT）中，将 GFF 注射到去核绵羊中期 II 期卵母细胞的卵质中，然后在体外培养非激活重构胚胎，使供体核能够暴露于卵质一段时间。随后，在第二阶段核移植中，将 FSNT 衍生的非激活重构胚胎离心，然后将供体核转移到另一个新去核的绵羊卵母细胞中。与一阶段核移植相比，两阶段核移植可以显著提高山羊-绵羊种间克隆胚胎的囊胚率，这一结果表明长时间暴露于异种卵质有利于供体核的重编程。两阶段核移植程序有两个优点，一是供体核可以长时间暴露于卵质中，二是可以解决卵母细胞老化的问题。
