This study was conducted to determine the effect of microinjection of a single blastomere from in vitro fertilization (IVF)-derived eight-cell embryo into eight-cell cloned embryos harboring the gene encoding recombinant human lactoferrin (rhLF), GFP, and NEO markers in bovine. The reconstructed chimeric embryos were assessed for their development to blastocyst, or to term after transfer, and tissues of offspring were evaluated by polymerase chain reaction (PCR) for the presence of nuclear transfer (NT)-derived transgenic cells, and the cloned embryos without microinjection were used as controls. The chimeric embryos showed slightly higher blastocyst rate than that for controls. The single IVF-derived blastomere appeared to preferential contribute to inner cell mass (ICM) in the chiemric blastocysts. After transfer, the rates of development of chimeric embryos to day 60, to term, and to weaning were significantly higher than those of controls. Sixty-three chimeric blastocysts were transferred and 11 calves were born: 7 calves of them were dead, and the remaining 4 calves are apparently normal and healthy. Most of the tissues collected from dead fetus were transgenic, whereas NT-derived transgenic cells were not detected in some tissues of the living calves. Our results indicated that a single blastomere from IVF-derived eight-cell embryo improves the in vivo developmental potential of transgenic cloned eight-cell embryos in bovine; however, the single IVF-derived blastomere appeares to be better able to populate the ICM and many tissues of offspring than NT-derived blastomeres.
Get full access to this article
View all access options for this article.
References
1.
BeddingtonR.S., RobertsonE.J.1989. An assessment of the developmental potential of embryonic stem cells in the midgestation mouse embryo. Development, 105:733–737.
2.
BoedionoA., SuzukiT., LiL.Y.et al.1999. Offspring born from chimeras reconstructed from parthenogenetic and in vitro fertilized bovine embryos. Mol. Reprod. Dev., 53:159–170.
3.
BoianiM., EckardtS., LeuN.A.et al.2003. Pluripotency deficit in clones overcome by clone–clone aggregation: epigenetic complementation?EMBO J., 22:5304–5312.
4.
ChoS.K., KimJ.H., ParkJ.Y.et al.2007. Serial cloning of pigs by somatic cell nuclear transfer: restoration of phenotypic normality during serial cloning. Dev. Dyn., 236:3369–3382.
5.
CibelliJ.B., SticeS.L., GoluekeP.J.et al.1998. Transgenic bovine chimeric offspring produced from somatic cell-derived stem-like cells. Nat. Biotechnol., 16:642–646.
6.
CibelliJ.B., CampbellK.H., SeidelG.E.et al.2002. The health profile of cloned animals. Nat. Biotechnol., 20:13–14.
7.
ClarkeH., VarmuzaS., PrideauxV.et al.1988. The development potential of parthenogenetically derived cells in chimeric mouse embryos: implications for action of imprinted genes. Development, 104:175–182.
8.
ConstantF., GuillomotM., HeymanY.et al.2006. Large offspring or large placenta syndrome? Morphometric analysis of late gestation bovine placentomes from somatic nuclear transfer pregnancies complicated by hydrallantois. Biol. Reprod., 75:122–130.
9.
DaliriM., RaoK.B., KaurG.et al.1999. Expression of growth factor ligand and receptor genes in preimplantation stage water buffalo (Bubalus bubalis) embryos and oviduct epithelial cells. J. Reprod. Fertil., 117:61–70.
10.
DeanW., SantosF., StojkovicM.et al.2001. Conservation of methylation reprogramming in mammalian development: aberrant reprogramming in cloned embryos. Proc. Natl. Acad. Sci. USA, 98:13734–13738.
11.
DechiaraT.M., PoueymirouW.T., AuerbachW.et al.2009. VelociMouse: fully ES cell-derived F0-generation mice obtained from the injection of ES cells into eight-cell-stage embryos. Methods Mol. Biol., 530:311–324.
12.
DucibellaT., AndersonE.1975. Cell shape and membrane changes in the eight-cell mouse embryo: prerequisites for morphogenesis of the blastocyst. Dev. Biol., 47:45.
13.
EnrightB.P., SungL.Y., ChangC.C.et al.2005. Methylation and acetylation characteristics of cloned bovine embryos from donor cells treated with 5-aza-2′-deoxycytidine. Biol. Reprod., 72:944–948.
14.
FreiR.E., SchultzG.A., ChurchR.B.1989. Qualitative and quantitative changes in protein synthesis occur at the 8–16-cell stage of embryogenesis in the cow. J. Reprod. Fertil., 86:637–641.
15.
GibbonsJ., AratS., RzucidloJ.et al.2002. Enhanced survivability of cloned calves derived from roscovitine-treated adult somatic cells. Biol. Reprod., 66:895–900.
16.
GongG., DaiY., FanB.et al.2004. Production of transgenic blastocyst by nuclear transfer from different types of somatic cells in cattle. Sci. China C Life Sci., 47:183–189.
17.
HallV.J., RuddockN.T., CooneyM.A.et al.2006. Production of a cloned calf using zona-free serial nuclear transfer. Theriogenology, 65:424–440.
18.
HiendlederS., MundC., ReichenbachH.D.et al.2004. Tissue-specific elevated genomic cytosine methylation levels are associated with an overgrowth phenotype of bovine fetuses derived by in vitro techniques. Biol. Reprod., 71:217–223.
19.
HillJ.R., RousselA.J., CibelliJ.B.et al.1999. Clinical and pathologic features of cloned transgenic calves and fetuses (13 case studies)Theriogenology, 51:1451–1465.
20.
HuangJ., DengK., WuH.et al.2008. Efficient production of mice from embryonic stem cells injected into four- or eight-cell embryos by piezo micromanipulation. Stem Cells, 26:1883–1890.
21.
IwasakiS., YoshibaN., UshijimaH.et al.1990. Morphology and proportion of inner cell mass of bovine blastocysts fertilized in vitro and in vivo. J. Reprod. Fertil., 90:279–284.
22.
KasinathanP., KnottJ.G., WangZ.et al.2001. Production of calves from G1 fibroblasts. Nat. Biotechnol., 19:1176–1178.
23.
KimberS.J., SuraniM.A., BartonS.C.1982. Interactions of blastomeres suggest changes in cell surface adhesiveness during the formation of inner cell mass and trophectoderm in the preimplantation mouse embryo. J. Embryol. Exp. Morphol., 70:133–152.
24.
KooD.B., KangY.K., ChoiY.H.et al.2002. Aberrant allocations of inner cell mass and trophectoderm cells in bovine nuclear transfer blastocysts. Biol. Reprod., 67:487–492.
25.
KoyamaH., SuzukiH., YangX.et al.1994. Analysis of polarity of bovine and rabbit embryos by scanning electron microscopy. Biol. Reprod., 50:163–170.
26.
Misica-TurnerP.M., ObackF.C., EichenlaubM.et al.2007. Aggregating embryonic but not somatic nuclear transfer embryos increases cloning efficiency in cattle. Biol. Reprod., 76:268–278.
27.
NagyA., SassM., MarkkulaM.1989. Systematic non-uniform distribution of parthenogenetic cells in adult mouse chimaeras. Development, 106:321–324.
28.
NiemannH., TianX.C., KingW.A.et al.2008. Epigenetic reprogramming in embryonic and foetal development upon somatic cell nuclear transfer cloning. Reproduction, 135:151–163.
29.
PalmieriC., LoiP., PtakG.et al.2008. Review paper: a review of the pathology of abnormal placentae of somatic cell nuclear transfer clone pregnancies in cattle, sheep, and mice. Vet. Pathol., 45:865–880.
RhoG.J., KangT.Y., KochharH.P.et al.2001. Effect of blastomere sex and fluorescent labelling on the development of bovine chimeric embryos reconstituted at the four-cell stage. Mol. Reprod. Dev., 60:202–207.
32.
SaburiS., AzumaS., SatoE.et al.1997. Developmental fate of single embryonic stem cells microinjected into 8-cell-stage mouse embryos. Differentiation, 62:1–11.
33.
ShiW., HaafT.2002. Aberrant methylation patterns at the two-cell stage as an indicator of early developmental failure. Mol. Reprod. Dev., 63:329–334.
34.
TarkowskiA.K., WroblewskaJ.1967. Development of blastomeres of mouse eggs isolated at the 4- and 8-cell stage. J. Embryol. Exp. Morphol., 18:155–180.
35.
Van SoomA., BoerjanM.L., BolsP.E.et al.1997. Timing of compaction and inner cell allocation in bovine embryos produced in vivo after superovulation. Biol. Reprod., 57:1041–1049.
36.
WatsonA.J., HoganA., HahnelA.et al.1992. Expression of growth factor ligand and receptor genes in the preimplantation bovine embryo. Mol. Reprod. Dev., 31:87–95.
37.
WellsK.D., PowellA.M.2000. Blastomeres from somatic cell nuclear transfer embryos are not allocated randomly in chimeric blastocysts. Cloning, 2:9–22.
38.
WilladsenS., JanzenR., McAlisterR.et al.1991. The viability of late morulae and blastocysts produced by nuclear transplantation in cattle. Theriogenology, 35:161–170.
39.
WilladsenS.M., PolgeC.1981. Attempts to produce monozygotic quadruplets in cattle by blastomere separation. Vet. Rec., 108:211–213.
WoodS.A., PascoeW.S., SchmidtC.et al.1993. Simple and efficient production of embryonic stem cell–embryo chimeras by coculture. Proc. Natl. Acad. Sci. USA, 90:4582–4585.
43.
YangF., HaoR., KesslerB.et al.2007. Rabbit somatic cell cloning: effects of donor cell type, histone acetylation status and chimeric embryo complementation. Reproduction, 133:219–230.
44.
YangP., WangJ., GongG.et al.2008. Cattle mammary bioreactor generated by a novel procedure of transgenic cloning for large-scale production of functional human lactoferrin. PLoS One, 3:e3453.
45.
YangX.Y., LiH., MaQ.W.et al.2006. Improved efficiency of bovine cloning by autologous somatic cell nuclear transfer. Reproduction, 132:733–739.
46.
ZhouW., XiangT., WalkerS.et al.2008. Aggregation of bovine cloned embryos at the four-cell stage stimulated gene expression and in vitro embryo development. Mol. Reprod. Dev., 75:1281–1289.
47.
ZiomekC.A., JohnsonM.H.1980. Cell surface interaction induces polarization of mouse 8-cell blastomeres at compaction. Cell, 21:935–942.
