Development of Human cloned Blastocysts Following Somatic Cell Nuclear Transfer (SCNT) with Adult Fibroblasts

¹ Stemagen, Corporation, La Jolla, California, USA
² The Reproductive Sciences Center, La Jolla, California, USA
³ Genesis Genetics Institute, LLC, Detroit, Michigan, USA
⁴ Stemagen, Corporation, La Jolla, California, USA; The Reproductive Sciences Center, La Jolla, California, USA

^* To whom correspondence should be addressed. E-mail: afrench{at}stemagen.com.

	Abstract

Nuclear transfer stem cells (NTSC) holds considerable promise in the field of regenerative medicine and cellbased drug discovery. In this study, a total of 29 oocytes were obtained from three young (20–24 y) reproductive egg donors who had been successful in previous cycles. These oocytes, deemed by intended parents to be in excess of their reproductive needs, were donated for research without financial compensation by both the egg donor and intended parents after receiving informed consent. All intended parents successfully achieved ongoing pregnancies with the oocytes retained for reproductive purposes. Mature oocytes, obtained within 2 h following transvaginal aspiration, were enucleated using one of two methods, extrusion or aspiration, after 45 min incubation in Cytochalasin B. Rates of oocyte lysis or degeneration did not differ between the two methods. Somatic cell nuclear transfer (SCNT) embryos were constructed using two established adult male fibroblast lines of normal karyotype. High rates of pronuclear formation (66%), early cleavage (47%) and blastocyst (23%) development were observed following incubation in standard IVF culture media. One cloned blastocyst was confirmed by DNA and mtDNA fingerprinting analyses and DNA fingerprinting of two other cloned blastocysts indicated they were also generated by SCNT. Blastocysts were also obtained from a limited number of parthenogenetically activated oocytes. This study demonstrates, for the first time, that SCNT can produce human blastocyst stage embryos using nuclei from a differentiated adult somatic cell, and provides new information on methods that may be needed for a higher level of efficiency for human therapeutic cloning.

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