Document Detail


Feasibility of producing porcine nuclear transfer embryos by using G2/M-stage fetal fibroblasts as donors.
MedLine Citation:
PMID:  11673275     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
The type of donor cell most suitable for producing cloned animals is one of the topics under debate in the field of nuclear transfer. To provide useful information to answer this question, G2/M- and G0/G1-stage fetal fibroblasts were used as donor cells for nuclear transfer. In vitro-matured oocytes derived from abattoir ovaries were used as recipient cytoplasts. In both groups, nuclear envelope breakdown and premature chromosome condensation were completed within 1-2 h after donor cells were injected into the cytoplasm of oocytes. Microtubules were organized around condensed chromosomes and formed a spindle within 1-1.5 h after activation. Decondensation of chromosomes could be seen within 2-4 h after activation. Reformation of the new nuclear envelope occurred 4-6 h after activation and was followed by nuclear swelling and formation of a pronucleus-like structure (PN) 8-12 h after activation. Most (80.6%) of the reconstructed oocytes derived from G2/M cells extruded polar body-like structures (PB). However, a much lower frequency of PB (21.7%) was observed in the reconstructed oocytes derived from G0/G1 donors. A variety of PN and PB combinations were observed in reconstructed oocytes derived from G2/M-stage donors, including 1PN+0PB, 1PN+1PB, 1PN+2PB, 2PN+0PB, 2PN+1PB, 2PN+2PB, and 3PN+1PB. Chromosomes of most embryos (10/13) derived from G2/M stage were diploid. The percentage of cleavage and blastocysts and the average nuclear number of blastocysts in the G2/M and G0/G1 groups were not different. These results demonstrate that the G2/M stage can be morphologically remodeled by cytoplasm of MII oocytes in pigs. To maintain normal ploidy, the extra chromosomes derived from G2/M-stage cells could be expelled by oocytes as a second polar body. G2/M-stage fibroblast nuclei could direct reconstructed embryos to develop to the blastocyst stage.
Authors:
L Lai; T Tao; Z Macháty; B Kühholzer; Q Y Sun; K W Park; B N Day; R S Prather
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Biology of reproduction     Volume:  65     ISSN:  0006-3363     ISO Abbreviation:  Biol. Reprod.     Publication Date:  2001 Nov 
Date Detail:
Created Date:  2001-10-23     Completed Date:  2002-01-22     Revised Date:  2007-11-15    
Medline Journal Info:
Nlm Unique ID:  0207224     Medline TA:  Biol Reprod     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1558-64     Citation Subset:  IM    
Affiliation:
Department of Animal Sciences, University of Missouri, Columbia, Missouri 65211, USA.
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MeSH Terms
Descriptor/Qualifier:
Animals
Blastocyst / physiology
Cell Cycle*
Chromosomes / ultrastructure
Cloning, Organism / methods*
Colchicine / pharmacology
Culture Techniques
Electric Stimulation
Embryo, Mammalian / cytology
Female
Fibroblasts / ultrastructure*
Flow Cytometry
G2 Phase
Microtubules / drug effects,  ultrastructure
Mitosis
Nuclear Envelope / ultrastructure
Nuclear Transfer Techniques*
Oocytes / ultrastructure*
Ploidies
Pregnancy
Swine*
Grant Support
ID/Acronym/Agency:
RR 13438/RR/NCRR NIH HHS
Chemical
Reg. No./Substance:
64-86-8/Colchicine

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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