title: Effect of demecolcine-assisted enucleation on the MPF level and cyclin B1 distribution in porcine oocytes.
abstract: Demecolcine (DEM) treatment of oocytes induces formation of a membrane protrusion containing a mass of condensed maternal chromosomes, which can be removed with minimal damage prior to somatic cell nuclear transfer (SCNT). However, the effect of this method on the distribution of maturation-promoting factor (MPF) in porcine oocytes has not been reported. Here, the level of MPF and the distribution of cyclin B1 were assessed in porcine oocytes following DEM treatment. In addition, the efficiencies of DEM-assisted and mechanical enucleation were compared, as were the development (in vitro and in vivo) of these oocytes following SCNT. MPF was uniformly distributed in oocytes that had been treated with 0.4 μg/ml DEM for 1 h. Immunofluorescence microscopy showed that in untreated oocytes, cyclin B1, the regulatory subunit of MPF, accumulated around the spindle, and was lowly detected in the cytoplasm. DEM treatment disrupted spindle microtubules, induced chromosome condensation, and reduced the level of cyclin B1 in the nuclear region. Cyclin B1 was uniformly distributed in DEM-treated oocytes and the level of MPF was increased. The potential of embryos generated from DEM-treated oocytes to develop in vivo was significantly greater than that of embryos generated from mechanically enucleated oocytes. This is the first study to report the effects of DEM-assisted enucleation of porcine oocytes on the distribution of cyclin B1. MPF in mature oocytes is important for the development of reconstructed embryos and for efficient SCNT.
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Suo
authors.initials: S
authors.lastname: Li
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Jin-Dan
authors.initials: JD
authors.lastname: Kang
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Jun-Xue
authors.initials: JX
authors.lastname: Jin
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Hong
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Hai-Ying
authors.initials: HY
authors.lastname: Zhu
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Long
authors.initials: L
authors.lastname: Jin
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Qing-Shan
authors.initials: QS
authors.lastname: Gao
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Chang-Guo
authors.initials: CG
authors.lastname: Yan
authors.affiliation: Department of veterinary medicine, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Cheng-Du
authors.initials: CD
authors.lastname: Cui
authors.affiliation: Department of veterinary medicine, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Wen-Xue
authors.initials: WX
authors.lastname: Li
authors.affiliation: Department of Animal Science, College of Agriculture, Yanbian University, Yanji, China.
authors.forename: Xi-Jun
authors.initials: XJ
authors.lastname: Yin
mesh_terms: D000818:Animals; D056744:Cyclin B1 / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D003703:Demecolcine / Q000737:chemistry*; D004423:Ear; D004624:Embryo Transfer; D005347:Fibroblasts / Q000166:cytology; D018507:Gene Expression Regulation, Developmental; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000187:drug effects; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000166:cytology / Q000378:metabolism*; D008941:Spindle Apparatus / Q000187:drug effects; D013552:Swine; D013556:Swine, Miniature; D050257:Tubulin Modulators / Q000737:chemistry*
chemical_list: D056744:Cyclin B1; D050257:Tubulin Modulators; D003703:Demecolcine