title: Germ-cell culture conditions facilitate the production of mouse embryonic stem cells.
abstract: The derivation of embryonic stem-cell (ESC) lines from blastocysts is a very inefficient process. Murine ESCs are thought to arise from epiblast cells that are already predisposed to a primordial-germ-cell fate. During the process of ESC derivation from B6D2 F1 hybrid mice, if we first culture the embryo from the two-cell stage in medium supplemented with LIF, we improve the quality of the blastocyst. When the blastocyst is then cultured in a germ-line stem-cell culture medium (GSCm), we are able to more efficiently (28.3%) obtain quality ESC lines that have a normal karyotype, proper degree of chimerism, and exhibit germ-line transmission when microinjected into blastocysts. Although germ-cell-specific genes were expressed in all culture medium conditions, GSCm did not shift the transcriptome towards germ-cell specification. A correlation was further observed between ESC derivation efficiency and the expression of some imprinted genes and retrotransposable elements. In conclusion, the combination of LIF supplementation followed by culture in GSCm establishes a higher efficiency method for ESC derivation.
authors.affiliation: Departamento de Reproducción Animal, INIA, Madrid, Spain.
authors.forename: Priscila
authors.initials: P
authors.lastname: Ramos-Ibeas
authors.forename: Eva
authors.initials: E
authors.lastname: Pericuesta
authors.forename: Raúl
authors.initials: R
authors.lastname: Fernández-González
authors.forename: Alfonso
authors.initials: A
authors.lastname: Gutiérrez-Adán
authors.forename: Miguel Ángel
authors.initials: MÁ
authors.lastname: Ramírez
mesh_terms: D000818:Animals; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D001755:Blastocyst / Q000166:cytology*; D018929:Cell Culture Techniques / Q000379:methods*; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005260:Female; D018507:Gene Expression Regulation, Developmental; D005819:Genetic Markers / Q000235:genetics; D018392:Genomic Imprinting; D008297:Male; D051379:Mice; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D062085:RNA, Long Noncoding / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; D005819:Genetic Markers; C422882:MEG3 non-coding RNA, mouse; D062085:RNA, Long Noncoding