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Epiblast isolation by a new four stage method (peeling) from whole bovine cloned blastocysts
Author(s) -
Kwon Dae Kee,
Hong So Gun,
Park Hee Jung,
Kang Jung Taek,
Koo Ok Jae,
Lee Byeong Chun
Publication year - 2009
Publication title -
cell biology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 77
eISSN - 1095-8355
pISSN - 1065-6995
DOI - 10.1016/j.cellbi.2008.12.003
Subject(s) - epiblast , blastocyst , endoderm , inner cell mass , biology , trophoblast , embryo , zona pellucida , microbiology and biotechnology , embryonic stem cell , genetics , embryogenesis , oocyte , gastrulation , fetus , gene , placenta , pregnancy
We have established a new 4 stage epiblast isolation method from whole bovine cloned blastocysts without using immunosurgery. The new “peeling” method consists of dissolution of the zona pellucida (first stage), elimination of mural trophoblast (second stage), isolation of primitive endoderm and epiblast from polar trophoblast (third stage), and isolation of epiblast from primitive endoderm (fourth stage). The bovine cloned blastocyst consists of 4 different types of cells showing abundant alkaline phosphatase activity. The epiblast origin of isolated cells was confirmed by in vitro differentiation of isolated cells to tubulin β3‐positive neurons and by embryoid body formation. The bovine cloned blastocyst origin of isolated epiblasts was confirmed by microsatellite analysis and mitochondrial DNA sequencing analysis. This new method might accelerate establishment of somatic cell nuclear transfer derived embryonic stem cell lines from bovine and other mammals.