title: Regulation of DNA synthesis at the first cell cycle in the sea urchin in vivo.
abstract: Using fluorescent and non-fluorescent recombinant proteins has proved to be a very successful technique for following postfertilization events, in both male and female pronuclei during the first cell cycle of sea urchin in vivo. Proteins and dyes are introduced by microinjection into the unfertilized egg, and their function can be monitored by fluorescence or confocal/two-photon (2P) and transmitted light microscopy after insemination. Here, we describe expression and purification of GFP/RFP-tagged proteins involved in regulation of DNA replication. We also explain the techniques used to introduce recombinant proteins and fluorescent tubulin into sea urchin eggs and embryos.
authors.affiliation: Institute of Cell and Molecular Biosciences, Faculty of Medical Sciences, The Medical School, Newcastle University, Newcastle upon Tyne, UK.
authors.forename: Jolanta
authors.initials: J
authors.lastname: Kisielewska
authors.forename: Michael
authors.initials: M
authors.lastname: Whitaker
mesh_terms: D000818:Animals; D002453:Cell Cycle; D002467:Cell Nucleus / Q000378:metabolism; D004261:DNA Replication; D004926:Escherichia coli; D005260:Female; D005306:Fertilization; D005456:Fluorescent Dyes / Q000737:chemistry / Q000302:isolation & purification; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D008297:Male; D008845:Microinjections; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D010063:Ovum / Q000166:cytology; D018809:Proliferating Cell Nuclear Antigen / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012617:Sea Urchins / Q000166:cytology* / Q000502:physiology; D013194:Staining and Labeling; D014404:Tubulin / Q000737:chemistry
chemical_list: D005456:Fluorescent Dyes; D018809:Proliferating Cell Nuclear Antigen; D014404:Tubulin; D049452:Green Fluorescent Proteins