title: Porcine Cloned Embryos Reconstructed with the Cell Nuclei of Tetraploid M-phase Fibroblast Cells Can Restore Normal Diploidy at the Blastocyst Stage.
abstract: The cell cycle of donor cells as a major factor that affects cloning efficiency remains debatable. G2/M phase cells as a donor can successfully produce cloned animals, but a minimal amount is known regarding nuclear remodeling events. In this study, porcine fetal fibroblasts (PFFs) were carefully synchronized at G1 or M phase as donor cells. Most of the cloned embryos reconstructed from PFFs at G1 (G1-embryos) or M (M-embryos) phase formed a pronucleus-like nucleus (PN) within 6-h post fusion (hpf), but the M-embryos formed PN earlier than the G1-embryos did. Moreover, 77.4% of the M-embryos formed two PNs, whereas the G1-embryos formed a single PN. The rate of extrusion of polar body-like structures by the M-embryos was significantly lower than that extruded by the G1-embryos (26.3% vs. 37.1%, P < 0.05), and DNA synthesis in most embryos in both groups was initiated at 9-12 hpf. Most of the M-embryos were octoploid before the first cleavage. Furthermore, 81.25% of the blastomeres of blastocysts developed from the M-embryos showed abnormal ploidy compared with those developed from the G1-embryos (22.55%). However, some of the blastomeres remained diploid in all the M-embryos tested. A portion of the blastomeres restored normal diploidy in some of the M-embryos at the blastocyst stage. This finding provides an explanation for M-embryos developing to term.
authors.affiliation: a College of Life Science , Northeast Forestry University , Harbin , P. R. China.
authors.forename: Q
authors.initials: Q
authors.lastname: Zhao
authors.affiliation: a College of Life Science , Northeast Forestry University , Harbin , P. R. China.
authors.forename: Y G
authors.initials: YG
authors.lastname: Qiu
authors.affiliation: b Department of Cardiology , The 2nd Affiliated Hospital of Harbin Medical University, The Key Laboratory of Myocardial Ischemia, Chinese Ministry of Education , Harbin , P. R. China.
authors.forename: J T
authors.initials: JT
authors.lastname: Tian
authors.affiliation: a College of Life Science , Northeast Forestry University , Harbin , P. R. China.
authors.forename: C S
authors.initials: CS
authors.lastname: Wang
authors.affiliation: a College of Life Science , Northeast Forestry University , Harbin , P. R. China.
authors.forename: T Z
authors.initials: TZ
authors.lastname: An
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology*; D002453:Cell Cycle / Q000235:genetics; D065150:Cellular Reprogramming / Q000235:genetics; D019976:Cloning, Organism / Q000379:methods*; D004171:Diploidy; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000166:cytology*; D059785:Karyotype; D013552:Swine; D057891:Tetraploidy
keywords: Blastocyst stage; G1/M phase; cloned porcine; diploidy; embryo