组织工程研究中的胚胎干细胞问题

胚胎干细胞研究是组织工程基本科学问题中一个重要的组成部分。本文回顾了近年来胚胎干细胞领域的研究进程,尤其是人体胚胎干细胞研究的最新成果;提出了组织工程中“治疗性克隆”的战略目标。并且确立了未来人体胚胎干细胞研究解决组织工程种子细胞问题的具体步骤;指出了每项具体研究中所要解决的关键问题。     

Somatic cell bovine cloning: Effect of donor cell and recipients

Nuclear transfer (NT) is an efficient technique for assessing the developmental potential of a nucleus and for analyzing the interactions between the donor nucleus and the recipient cytoplasm. In amphibians, thought nuclei of adult kerationocytes support development to the juvenile, tadpole stage, no development to the adult stage was re- ported[1], leaving open the question of whether a different- tiated adult nucleus can be fully reprogrammed. The first cloned offspring developed from differ…     

从蝾螈到克隆猴:核移植研究90周年记

从第一例核移植动物到第一例体细胞核移植灵长类动物,核移植研究恰好迎来90周年。在这近百年的研究历程中,核移植研究不仅回答了一系列重大科学问题,且在许多方面已投入到实际应用。文章详述了核移植研究的发展历程、实际应用及其发展现状。     

电脉冲激活猪卵母细胞的研究

通过对猪卵母细胞的电激活进行系统研究，结果表明：对于体外成熟母卵细胞，电场强度为１２５０Ｖ／ｃｍ，脉冲时程为４０，６０μｓ时，效果较好，一次脉冲激活率可达８５．３％和８７．１％，而对于体内成熟卵母细胞，电场强度为１５００Ｖ／ｃｍ，脉冲时间为４０，６０μｓ时，效果了，一次脉冲激活率达８５．７％和８８．２％；     

兔ES样细胞系的建立及其特性分析

报道从２３７枚家兔胚胎中建成７个可连续传代的ＥＳ样细胞系。建系条件为，使用小鼠原始胚胎成纤维细胞（ＭＥ）作饲养层，以含１０％胎年血清和１０％兔血清的ＤＭＥＭ／Ｆ１２为培养基，添加白血病抑制因子（ＬＩＦ）或上皮生长因子（ＥＧＦ），胚龄为９０，９６ｈ。该细胞系的细胞。在许多方面类似于小鼠ＥＳ细胞，具干细胞的形态特征，呈集落型生长，可连续传代并保持其形态特征，具有一定的自发分化和诱导分化的能力，悬浮培养     

卵母细胞的卵龄对猪核移植效率的影响

对核移植中，猪卵母细胞的卵龄对核移植效率的影响进行了研究。结果表明：体外成熟培养４８ｈ的猪卵母细胞最适合作为核受体其激活率达７８．３％，并无裂解：去核率高达９０％；在作为核受体进行核移植时，融合率为７５％，重构胚胎的卵裂率为６４．６％，卵龄进一步增大，激活率略有增加，但去核率，融合率和卵裂率均下降。     

乳腺组织特异表达h—LTF转基因山羊研究

该研究以改构的hLTF基因为目的基因,利用基因打靶技术将其定位敲入到山羊胎儿成纤维细胞的β-casein基因座,检测阳性的基因打靶克隆扩增后用于体细胞核移植,核移植胚胎体外培养发育到2-细胞期后进行胚胎移植。结果表明,通过研究首次建立了山羊胎儿成纤维细胞和乳腺上皮细胞乳蛋白基因座高效基因打靶的技术体系以及基因打靶体细胞核移植技术体系。经过G418药物抗性筛选后获得了稳定整合人lactoferrin基因的转基因阳性细胞克隆14个,体内成熟卵母细胞作受体的核移植囊胚率为64．8％,体外成熟卵母细胞作受体的核移植囊胚率为51．7％。利用DMSO-SMGT技术制备转基因动物高阳性率为56．5％,获得了转基因小鼠8只、转基因兔155只、转基因山羊2只。在转基因后代的乳腺里,外源基因得到了表达,并且具有生物活性,建立了DMSO-SMGT技术体系。     

Effects of different nuclear transfer and activation methods on the development of mouse somatic cell cloned embryos

A group of adult somatic cell cloned mice were obtained by using cumulus cells as nuclei donor cells. To study the effect of different nuclear transfer (NT) and activation methods on the development of mouse cloned embryos, embryos were reconstructed using two traditional NT methods (electrofusion and direct injection) and four activation treatments (electric pulse, ethanol, SrCl2 and electric pulse combined with SrCl2). The data showed that the efficiency of reconstruction using the direct injection method is significantly higher (90.7%) than that of the electrofusion method (49.7%). Parthenogenetic embryos can develop to blastocyst stage with three activation conditions, including ethanol, electric pulse and SrCl2; however, the rates of development to blastocyst after ethanol and electric pulse acti-vation (52.4%, 54.2%) are significantly lower than after SrCl2 activation (76.9%). Treatment of embryos for 6 h with 10 mmol/L SrCl2 was found to be the best condition for activation of parthenogenetic as well as reconstructed embryos. By contrast, reconstructed embryos failed to develop to blastocyst stage after being activated by ethanol. The use of either injection or electrofusion for embryo reconstruction affected the pre-implantation development. However, after transfer in pseudopregnant mice, cloned mice were obtained from both methods.     

Expression of chromatin modification genes in organs of cloned cattle that died within hours after birth

Several mammalian species including sheep[1,2], bo- vine[3], goats[4], mice[5], pigs[6], cat[7] and rabbits[8], have been successfully cloned by somatic nuclear transfer (NT), but the efficiency of this process is poor, with only a small proportion of the…