Baguisi A, Behboodi E, Melican D T, Pollock J S, Destrempes M M, Cammuso C, Williams J L, Nims S D, Porter C A, Midura P, Palacios M J, Ayres S L, Denniston R S, Hayes M L, Ziomek C A, Meade H M, Godke R A, Gavin W G, Overström E W, Echelard Y
Department of Biomedical Sciences, Tufts University School of Veterinary Medicine, North Grafton, MA 01356, USA.
Nat Biotechnol. 1999 May;17(5):456-61. doi: 10.1038/8632.
In this study, we demonstrate the production of transgenic goats by nuclear transfer of fetal somatic cells. Donor karyoplasts were obtained from a primary fetal somatic cell line derived from a 40-day transgenic female fetus produced by artificial insemination of a nontransgenic adult female with semen from a transgenic male. Live offspring were produced with two nuclear transfer procedures. In one protocol, oocytes at the arrested metaphase II stage were enucleated, electrofused with donor somatic cells, and simultaneously activated. In the second protocol, activated in vivo oocytes were enucleated at the telophase II stage, electrofused with donor somatic cells, and simultaneously activated a second time to induce genome reactivation. Three healthy identical female offspring were born. Genotypic analyses confirmed that all cloned offspring were derived from the donor cell line. Analysis of the milk of one of the transgenic cloned animals showed high-level production of human antithrombin III, similar to the parental transgenic line.
在本研究中,我们展示了通过胎儿体细胞的核移植来生产转基因山羊。供体核质体取自一个原代胎儿体细胞系,该细胞系源自一只40天的转基因雌性胎儿,此胎儿是通过用转基因雄性的精液对非转基因成年雌性进行人工授精产生的。通过两种核移植程序产生了活体后代。在一种方案中,将处于中期II阻滞阶段的卵母细胞去核,与供体体细胞进行电融合,并同时激活。在第二种方案中,将体内激活的卵母细胞在末期II阶段去核,与供体体细胞进行电融合,并再次同时激活以诱导基因组重新激活。三只健康的同卵雌性后代出生。基因型分析证实所有克隆后代均源自供体细胞系。对其中一只转基因克隆动物的乳汁分析表明,其人类抗凝血酶III产量很高,与亲代转基因系相似。