Derivation and characterization of haploid embryonic stem cell cultures in medaka fish.
Embryonic stem (ES) cells are pluripotent cells capable of differentiation into various cell types. Haploid ES cells elegantly combine the advantages of haploidy and pluripotency and offer a unique in vitro system for genetic analyses of molecular, cellular and developmental events in various cell lineages. Our recent success in generating haploid ES cell lines from gynogenetic embryos of the medaka fish suggests that haploidy can support ES cell derivation and maintenance in a vertebrate. In this study, we present a step-by-step protocol for derivation and characterization of medaka haploid ES cells. We have used this procedure to produce three haploid ES cell lines from five primary cultures. It takes about 15 weeks to generate stable cultures, 5-8 weeks to obtain pure haploid cells and 5-6 weeks to characterize ES cells in vitro and in vivo.
Yi
Meisheng
M
Department of Biological Sciences, National University of Singapore, Singapore.
Hong
Ni
N
Hong
Yunhan
Y
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation; D049108:Cell Enlargement; D002460:Cell Line; D049109:Cell Proliferation; D015925:Cryopreservation; D003470:Culture Media; D004625:Embryo, Nonmammalian / Q000166:cytology; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005819:Genetic Markers; D049452:Green Fluorescent Proteins / Q000032:analysis; D006238:Haploidy; D009990:Oryzias / Q000196:embryology* / Q000235:genetics
D003470:Culture Media; D005819:Genetic Markers; D049452:Green Fluorescent Proteins