Somatic cell nuclear transfer (SCNT) is the process whereby the nucleus from an adult cell is transferred into a previously enucleated oocyte, followed by oocyte activation and the ultimate production of a clone of the original source (adult) cell.
SCNT is an inefficient process and substantial loss occurs throughout development. The precise mechanisms that are involved in these losses remain poorly understood.
Nobody has yet provided scientifically peer-reviewed proof of successful SCNT in humans or non-human primates. In rhesus macaques, this failure is the result of a difference in mitotic spindle organization.
A range of abnormalities have been shown in species in which cloned animals have been successfully produced; some defects are similar between species, but others are not. These defects include placental abnormalities, and kidney, lung and heart pathology.
Genetic and epigenetic effects are involved in cloning failures. DNA methylation, genomic imprinting, gene reprogramming, chromatin structure, X-chromosome inactivation and telomere lengths have all been shown to be influenced by cloning.
Interpreting the evidence from abnormal clones (both phenotypic, genetic and epigenetic data) is not straightforward. Some argue that the abnormalities reflect problems that are seen in association with natural reproduction (only at a higher level); others argue that the problems are unique and the mechanisms are poorly understood.
Experiments must be designed to examine the molecular, developmental and pathological issues in parallel. Such studies must be prospective and involve single-parameter changes, ideally in a range of species.
Although scientists should not condone human reproductive cloning, it is important to address the safety issues of cloning as it pertains to non-reproductive (therapeutic) cloning.
There are continued claims of attempts to clone humans using nuclear transfer, despite the serious problems that have been encountered in cloning other mammals. It is known that epigenetic and genetic mechanisms are involved in clone failure, but we still do not know exactly how. Human reproductive cloning is unethical, but the production of cells from cloned embryos could offer many potential benefits. So, can human cloning be made safe?
This is a preview of subscription content, access via your institution
Subscribe to this journal
Receive 12 print issues and online access
$189.00 per year
only $15.75 per issue
Rent or buy this article
Get just this article for as long as you need it
Prices may be subject to local taxes which are calculated during checkout
Wilmut, I., Schnieke, A. E., McWhir, J., Kind, A. J. & Campbell, K. H. S. Viable offspring derived from fetal and adult mammalian cells. Nature 385, 810–813 (1997). This paper describes the production of 'Dolly' the sheep — the first mammal to be successfully cloned from a differentiated adult (mammary) cell.
Zavos, P. M. Human reproductive cloning: the time is near. Reprod. Biomed. Online 6, 397–398 (2003).
Meng, L., Ely, J. J., Stouffer, R. L. & Wolf, D. P. Rhesus monkeys produced by nuclear transfer. Biol. Reprod. 57, 454–459 (1997).
Wolf, D. P., Meng, L., Ouhibi, N. & Zelinski-Wooten, M. Nuclear transfer in the rhesus monkey: practical and basic implications. Biol. Reprod. 60, 199–204 (1999).
Simerly, C. et al. Molecular correlates of primate nuclear transfer failures. Science 300, 297–297 (2003). This article presents evidence for fundamental differences in the organization of the mitotic spindle in rhesus macaques, which results in the removal of crucial proteins during the enucleation process.
Chesne, P. et al. Cloned rabbits produced by nuclear transfer from adult somatic cells. Nature Biotechnol. 20, 366–369 (2002).
King, T. J. et al. Embryo development and establishment of pregnancy after embryo transfer in pigs: coping with limitations in the availability of viable embryos. Reproduction 123, 507–515 (2002).
De Sousa, P. A. et al. Somatic cell nuclear transfer in the pig: control of pronuclear formation and integration with improved methods for activation and maintenance of pregnancy. Biol. Reprod. 66, 642–650 (2002).
Woods, G. L. et al. A mule cloned from foetal cells by nuclear transfer. Science 301, 1063 (2003).
Zhou, Q. et al. Generation of fertile cloned rats using controlled timing of oocyte activation. Science 25 September 2003 (doi:10.1126/science.1088313). This was the first report of successful cloning of the rat by controlling oocyte activation.
Hayes, E. et al. Nuclear transfer of adult and genetically modified fetal cells of the rat. Physiol. Genomics 5, 193–203 (2001).
Young, L. E., Sinclair, K. D. & Wilmut, I. Large offspring syndrome in cattle and sheep. Rev. Reprod. 3, 155–163 (1998).
Reik, W. & Maher, E. R. Imprinting in clusters: lessons from Beckwith–Wiedemann syndrome. Trends Genet. 13, 330–334 (1997).
DeBaun, M. R., Niemitz, E. L. & Feinberg, A. P. Association of in vitro fertilization with Beckwith–Wiedemann syndrome and epigenetic alterations of LIT1 and H19. Am. J. Hum. Genet. 72, 156–160 (2003).
Hill, J. R. et al. Evidence for placental abnormality as the major cause of mortality in first-trimester somatic cell cloned bovine fetuses. Biol. Reprod. 63, 1787–1794 (2000).
De Sousa, P. A. et al. Evaluation of gestational deficiencies in cloned sheep fetuses and placentae. Biol. Reprod. 65, 23–30 (2001). References 15 and 16 document placental abnormalities in cloned sheep and cattle.
Tanaka, S. et al. Placentomegaly in cloned mouse concepti caused by expansion of the spongiotrophoblast layer. Biol. Reprod. 65, 1813–1821 (2001).
Booth, P. J. et al. Numerical chromosome errors in day 7 somatic nuclear transfer bovine blastocysts. Biol. Reprod. 68, 922–928 (2003).
Tamashiro, K. L. K. et al. Cloned mice have an obese phenotype not transmitted to their offspring. Nature Med. 8, 262–267 (2002). This paper shows that the increased body weight of cloned female mice reflects an increase in body fat as well as larger body size; this obese phenotype is not transmitted to offspring.
Santos, F. et al. Epigenetic marking correlates with developmental potential in cloned bovine preimplantation embryos. Curr. Biol. 19, 1118–1121 (2003). This paper reports histone and DNA hypermethylation in cloned bovine embryos and provides further evidence that the epigenotype of cloned embryos depends on the donor-cell type.
Li, E. Chromatin modification and epigenetic reprogramming in mammalian development. Nature Rev. Genet. 3, 662–673 (2002).
Selig, S., Ariel, M., Goitein, R., Marcus, M. & Cedar, H. Regulation of mouse satellite DNA-replication time. EMBO J. 7, 419–426 (1988).
Jaenisch, R. DNA methylation and imprinting: why bother? Trends Genet. 13, 323–329 (1997).
Barlow, D. P. Methylation and imprinting — from host defense to gene-regulation. Science 260, 309–310 (1993).
Li, E., Beard, C. & Jaenisch, R. Role for DNA methylation in genomic imprinting. Nature 366, 362–365 (1993).
Jones, P. L. & Wolffe, A. P. Relationships between chromatin organization and DNA methylation in determining gene expression. Seminars Cancer Biol. 9, 339–347 (1999).
Shemer, R. et al. Dynamic methylation adjustment and counting as part of imprinting mechanisms. Proc. Natl Acad. Sci. USA 93, 6371–6376 (1996).
Mayer, W., Niveleau, A., Walter, J., Fundele, R. & Haaf, T. Embryogenesis — demethylation of the zygotic paternal genome. Nature 403, 501–502 (2000).
Oswald, J. et al. Active demethylation of the paternal genome in the mouse zygote. Curr. Biol. 10, 475–478 (2000).
Rougier, N. et al. Chromosome methylation patterns during mammalian preimplantation development. Genes Dev. 12, 2108–2113 (1998).
Monk, M., Boubelik, M. & Lehnert, S. Temporal and regional changes in DNA methylation in the embryonic, extraembryonic and germ-cell lineages during mouse embryo development. Development 99, 371–382 (1987).
Dean, W. et al. Conservation of methylation reprogramming in mammalian development: aberrant reprogramming in cloned embryos. Proc. Natl Acad. Sci. USA 98, 13734–13738 (2001). This paper shows that epigenetic reprogramming occurs aberrantly in most cloned embryos.
Wilmut, I. et al. Somatic cell nuclear transfer. Nature 419, 583–586 (2002).
Kang, Y. K. et al. Aberrant methylation of donor genome in cloned bovine embryos. Nature Genet. 28, 173–177 (2001). These results show highly aberrant methylation patterns in various genomic regions of cloned embryos.
Bourc'his, D. et al. Delayed and incomplete reprogramming of chromosome methylation patterns in bovine cloned embryos. Curr. Biol. 11, 1542–1546 (2001).
Kang, Y. K. et al. Limited demethylation leaves mosaic-type methylation states in cloned bovine pre-implantation embryos. EMBO J. 21, 1092–1100 (2002).
Chung, Y. G., Ratnam, S., Chaillet, J. R. & Latham, K. E. Abnormal regulation of DNA methyltransferase expression in cloned mouse embryos. Biol. Reprod. 69, 146–153 (2003).
Kang, Y. K. et al. Typical demethylation events in cloned pig embryos — clues on species-specific differences in epigenetic reprogramming of a cloned donor genome. J. Biol. Chem. 276, 39980–39984 (2001).
Mann, M. R. W. et al. Disruption of imprinted gene methylation and expression in cloned preimplantation stage mouse embryos. Biol. Reprod. 69, 902–914 (2003). This study shows that epigenetic errors can arise early in clone development.
Barlow, D. P. Gametic imprinting in mammals. Science 270, 1610–1613 (1995).
Chavatte-Palmer, P. et al. Clinical, hormonal, and hematologic characteristics of bovine calves derived from nuclei from somatic cells. Biol. Reprod. 66, 1596–1603 (2002).
Rhind, S. et al. Cloned lambs: lessons from pathology. Nature Biotechnol. 21, 10–11 (2003).
Young, L. E. & Fairburn, H. R. Improving the safety of embryo technologies: possible role of genomic imprinting. Theriogenology 53, 627–648 (2000).
Kruip, T. A. M. & den Daas, J. H. G. In vitro produced and cloned embryos: effects on pregnancy, parturition and offspring. Theriogenology 47, 43–52 (1997).
Young, L. E. et al. Epigenetic change in IGF2R is associated with fetal overgrowth after sheep embryo culture. Nature Genet. 27, 153–154 (2001). This paper shows reduced fetal methylation and expression of ovine IGF2R in association with large offspring syndrome.
Humpherys, D. et al. Epigenetic instability in ES cells and cloned mice. Science 293, 95–97 (2001). This study reports epigenetic abnormalities in both ES cells and clones, with the further implication that even apparently normal cloned animals might have subtle abnormalities in gene expression.
Avner, P. & Heard, E. X-chromosome inactivation: counting, choice and initiation. Nature Rev. Genet. 2, 59–67 (2001).
Eggan, K. et al. X-chromosome inactivation in cloned mouse embryos. Science 290, 1578–1581 (2000).
Wrenzycki, C. et al. In vitro production and nuclear transfer affect dosage compensation of the X-linked gene transcripts G6PD, PGK, and Xist in preimplantation bovine embryos. Biol. Reprod. 66, 127–134 (2002).
Daniels, R., Hall, V. J., French, A. J., Korfiatis, N. A. & Trounson, A. O. Comparison of gene transcription in cloned bovine embryos produced by different nuclear transfer techniques. Mol. Reprod. Devel. 60, 281–288 (2001).
Boiani, M., Eckardt, S., Scholer, H. R. & McLaughlin, K. J. Oct4 distribution and level in mouse clones: consequences for pluripotency. Genes Dev. 16, 1209–1219 (2002).
Bortvin, A. et al. Incomplete reactivation of Oct4-related genes in mouse embryos cloned from somatic nuclei. Development 130, 1673–1680 (2003). References 51 and 52 emphasize the importance of Oct4 and related genes in development: variations contribute to embryonic lethality.
Pesce, M. & Scholer, H. R. Oct-4: gatekeeper in the beginnings of mammalian development. Stem Cells 19, 271–278 (2001).
Wrenzycki, C. et al. Nuclear transfer protocol affects messenger RNA expression patterns in cloned bovine blastocysts. Biol. Reprod. 65, 309–317 (2001).
Shiels, P. G. et al. Analysis of telomere lengths in cloned sheep. Nature 399, 316–317 (1999).
Wakayama, T. et al. Ageing — cloning of mice to six generations. Nature 407, 318–319 (2000).
Lanza, R. P. et al. Extension of cell life-span and telomere length in animals cloned from senescent somatic cells. Science 288, 665–669 (2000).
Tian, X. C., Xu, J. & Yang, X. Z. Normal telomere lengths found in cloned cattle. Nature Genet. 26, 272–273 (2000).
Betts, D. H. et al. Reprogramming of telomerase activity and rebuilding of telomere length in cloned cattle. Proc. Natl Acad. Sci. USA 98, 1077–1082 (2001).
Miyashita, N. et al. Remarkable differences in telomere lengths among cloned cattle derived from different cell types. Biol. Reprod. 66, 1649–1655 (2002). References 55–60 emphasize the variability in data on telomere lengths, which can be either shortened, normal or lengthened depending on the cell type that is used in the cloning procedure.
Daniels, R., Hall, V. & Trounson, A. O. Analysis of gene transcription in bovine nuclear transfer embryos reconstructed with granulosa cell nuclei. Biol. Reprod. 63, 1034–1040 (2000).
Cibelli, J. B., Campbell, K. H., Seidel, G. E., West, M. D. & Lanza, R. P. The health profile of cloned animals. Nature Biotechnol. 20, 13–14 (2002).
Lanza, R. P. et al. Cloned cattle can be healthy and normal. Science 294, 1893–1894 (2001).
Keefer, C. L. et al. Production of cloned goats after nuclear transfer using adult somatic cells. Biol. Reprod. 66, 199–203 (2002).
Winger, Q. A. et al. Genetic reprogramming of lactate dehydrogenase, citrate synthase, and phosphofructokinase mRNA in bovine nuclear transfer embryos produced using bovine fibroblast cell nuclei. Mol. Reprod. Devel. 56, 458–464 (2000).
Vanstekelenburghamers, A. E. P. et al. Stage-specific appearance of the mouse antigen Tec-3 in normal and nuclear transfer bovine embryos — reexpression after nuclear transfer. Mol. Reprod. Devel. 37, 27–33 (1994).
Koo, D. B. et al. Developmental potential and transgene expression of porcine nuclear transfer embryos using somatic cells. Mol. Reprod. Dev. 58, 15–21 (2001).
Zavos, P. Committee of scientists for safe and responsible therapeutic human cloning. [online], (cited 29 Sep. 2003), <http://reproductivecloning.net/articles/zavos.htm (2001).
Braude, P., Pickering, S., Flinter, F. & Ogilvie, C. M. Preimplantation genetic diagnosis. Nature Rev. Genet. 3, 941–953 (2002).
Niemann, H. & Wrenzycki, C. Alterations of expression of developmentally important genes in preimplantation bovine embryos by in vitro culture conditions: implications for subsequent development. Theriogenology 53, 21–34 (2000).
Cezar, G. G. et al. Genome-wide epigenetic alterations in cloned bovine fetuses. Biol. Reprod. 68, 1009–1014 (2003).
Humphreys, D. et al. Abnormal gene expression in cloned mice derived from embryonic stem cell and cumulus cell nuclei. Proc. Natl Acad. Sci. USA 99, 12889–12894 (2002).
Wrenzycki, C. et al. Effects of culture system and protein supplementation on mRNA expression in pre-implantation bovine embryos. Hum. Reprod. 16, 893–901 (2001).
Nagy, A., Rossant, J., Nagy, R., Abramownewerly, W. & Roder, J. C. Derivation of completely cell culture-derived mice from early-passage embryonic stem-cells. Proc. Natl Acad. Sci. USA 90, 8424–8428 (1993).
Brambrink, T. et al. Application of cDNA arrays to monitor mRNA profiles in single preimplantation mouse embryos. Biotechniques 33, 376–379 (2002).
Balog, R. P. et al. Parallel assessment of CpG methylation by two-color hybridization with oligonucleotide arrays. Anal. Biochem. 309, 301–310 (2002).
Pritchard, C. C., Hsu, L., Delrow, J. & Nelson, P. S. Project normal: defining normal variance in mouse gene expression. Proc. Natl Acad. Sci. USA 98, 13266–13271 (2001). The studies described in this paper helped to define the baseline level of variability in mouse gene expression and emphasize the importance of replicate microarray experiments.
Evans, W. E. & Relling, M. V. Pharmacogenomics: translating functional genomics into rational therapeutics. Science 286, 487–491 (1999).
Wakayama, T. et al. Differentiation of embryonic stem cell lines generated from adult somatic cells by nuclear transfer. Science 292, 740–743 (2001).
Rideout, W. M., Hochedlinger, K., Kyba, M., Daley, G. Q. & Jaenisch, R. Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell 109, 17–27 (2002).
Zwaka, T. P. & Thomson, J. A. Homologous recombination in human embryonic stem cells. Nature Biotechnol. 21, 319–321 (2003).
Hochedlinger, K. & Jaenisch, R. Mechanisms of disease: nuclear transplantation, embryonic stem cells, and the potential for cell therapy. N. Engl. J. Med. 349, 275–286 (2003).
Cibelli, J. B. et al. Parthenogenetic stem cells in nonhuman primates. Science 295, 819–819 (2002).
Munsie, M. J. et al. Isolation of pluripotent embryonic stem cells from reprogrammed adult mouse somatic cell nuclei. Curr. Biol. 10, 989–992 (2000).
Wakayama, T. & Yanagimachi, R. Cloning of male mice from adult tail-tip cells. Nature Genet. 22, 127–128 (1999).
Wakayama, T., Perry, A. C. F., Zuccotti, M., Johnson, K. R. & Yanagimachi, R. Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394, 369–374 (1998).
Plass, C. & Soloway, P. D. DNA methylation, imprinting and cancer. Eur. J. Hum. Genet. 10, 6–16 (2002).
Zakhartchenko, V. et al. Adult cloning in cattle: potential of nuclei from a permanent cell line and from primary cultures. Mol. Reprod. Devel. 54, 264–272 (1999).
Wells, D. N., Misica, P. M. & Tervit, H. R. Production of cloned calves following nuclear transfer with cultured adult mural granulosa cells. Biol. Reprod. 60, 996–1005 (1999).
Shiga, K., Fujita, T., Hirose, K., Sasae, Y. & Nagai, T. Production of calves by transfer of nuclei from cultured somatic cells obtained from Japanese black bulls. Theriogenology 52, 527–535 (1999).
Polejaeva, I. A. et al. Cloned pigs produced by nuclear transfer from adult somatic cells. Nature 407, 86–90 (2000).
Hill, J. R. et al. Clinical and pathologic features of cloned transgenic calves and fetuses (13 case studies). Theriogenology 51, 1451–1465 (1999).
Lai, L. X. et al. Production of α-1,3-galactosyltransferase knockout pigs by nuclear transfer coning. Science 295, 1089–1092 (2002).
Cibelli, J. B. et al. Cloned transgenic calves produced from nonquiescent fetal fibroblasts. Science 280, 1256–1258 (1998).
Ogonuki, O. et al. Early death of mice cloned from somatic cells. Nature Genet. 30, 253–254 (2002).
McCreath, K. J. et al. Production of gene-targeted sheep by nuclear transfer from cultured somatic cells. Nature 405, 1066–1069 (2000).
Kato, Y., Tani, T. & Tsunoda, Y. Cloning of calves from various somatic cell types of male and female adult, newborn and fetal cows. J. Reprod. Fertil. 120, 231–237 (2000).
Denning, C. et al. Deletion of the α(1,3) galactosyl transferase (GGTA1) gene and the prion protein (PrP) gene in sheep. Nature Biotechnol. 19, 559–562 (2001).
Renard, J. P. et al. Lymphoid hypoplasia and somatic cloning. Lancet 353, 1489–1491 (1999).
Kishi, M. et al. Nuclear transfer in cattle using colostrum-derived mammary gland epithelial, cells and ear-derived fibroblast cells. Theriogenology 54, 675–684 (2000).
Hill, J. R. et al. Development rates of male bovine nuclear transfer embryos derived from adult and fetal cells. Biol. Reprod. 62, 1135–1140 (2000).
Inui, A. Obesity — a chronic health problem in cloned mice? Trends Pharmacol. Sci. 24, 77–80 (2003).
We each acknowledge discussions with colleagues during the development of these ideas and are grateful for funding from the governments, charities and commercial agencies that are acknowledged in our research papers. We apologize to colleagues whose work we have been unable to cite, but limitations of space in many cases limit citations to reviews rather than the source papers.
- SOMATIC CELL NUCLEAR TRANSFER
(SCNT). The process by which the nucleus from an adult cell is transferred into a previously enucleated cell; the reconstructed oocyte is activated, which initiates subsequent development.
Literally means 'outside conventional genetics'; this term describes any heritable change in gene expression that is not caused by a change in DNA sequence.
- G0 PHASE
The resting phase of the cell cycle.
The removal of nuclear material.
A preimplantation embryo that contains a fluid-filled cavity (blastocoel), a focal cluster of cells from which the embryo will develop (inner cell mass) and peripheral trophoblast cells.
- LARGE-OFFSPRING SYNDROME
A syndrome that is characterized by fetal oversize and various organ pathologies.
An increase in overall body size and organ size relative to normal controls.
- INNER CELL MASS
(ICM). A small group of cells that are present in the blastocyst, which comprise undifferentiated cells.
A preimplantation embryo that consists of a solid cluster of cells.
Enlargement of the placenta beyond its normal size.
The outer structural layer of the placenta.
A single-celled fertilized embryo.
The state of those mechanisms that regulate gene expression and are transmitted to daughter cells.
- GENOMIC IMPRINTING
The epigenetic marking of a gene on the basis of parental origin, which results in monoallelic expression.
The outer epithelial layer of the blastocyst.
The period immediately before and after birth.
A short repeat sequence of DNA at the end of chromosomes, which both protects and ensures the complete replication of chromosome ends.
- GRANULOSA CELL
A somatic cell that is found in the ovarian follicles, which supports oocyte growth, maturation and ovulation.
- CUMULUS CELL
A cell that is found in the developing ovarian follicles.
Rights and permissions
About this article
Cite this article
Rhind, S., Taylor, J., De Sousa, P. et al. Human cloning: can it be made safe?. Nat Rev Genet 4, 855–864 (2003). https://doi.org/10.1038/nrg1205
This article is cited by
Screening somatic cell nuclear transfer parameters for generation of transgenic cloned cattle with intragenomic integration of additional gene copies that encode bovine adipocyte-type fatty acid-binding protein (A-FABP)
Molecular Biology Reports (2017)
Cells as irreducible wholes: the failure of mechanism and the possibility of an organicist revival
Biology & Philosophy (2013)
Advances in Reprogramming Somatic Cells to Induced Pluripotent Stem Cells
Stem Cell Reviews and Reports (2010)
Coplanar film electrodes facilitate bovine nuclear transfer cloning
Biomedical Microdevices (2009)
Fertilized eggs reprogram adult-cell genomes
Nature Reports Stem Cells (2007)