Embryonic Stem Cells: Methods and Protocols (Methods in Molecular Biology)

Embryonic stem (ES) cells have significant potential in basic studies designed to better understand how different cells and tissues in the body are formed, as well as for generating unlimited numbers of cells for transplantation, drug delivery, and drug testing. In Embryonic Stem Cells: Methods and Protocols, Kursad Turksen and a panel of international experts describe their most productive methods for using ES cells as in vitro developmental models for many cell and tissue types. Set out in step-by-step detail by the investigators who developed them, these protocols range widely from ES cell isolation, maintenance, and modulation of gene expression, to cutting-edge techniques that use cDNA arrays in gene expression analysis and phage display libraries. There are also advanced techniques for the generation of antibodies against very rare antigens and for the identification and characterization of proteins and protein interactions. Additional studies of the ES cell cycle and apoptosis, as well as protocols for the use of ES cells to generate diverse cell and tissue types, complete this collection of readily reproducible methods. Many of the techniques have already been shown to have tremendous utility with ES cells and their differentiated progeny. Authoritative and state-of-the-art, this unique first collection of protocols for the study of ES cells, Embryonic Stem Cells: Methods and Protocols, will prove an invaluable resource not only for those generally interested in cell and developmental biology, but also for those actively using, or planning to use, ES cells to study fate choices and specific lineages. Machine generated contents note: 1 Methods for the Isolation and Maintenance of Murine Embryonic -- Stem Cells -- Marsha L. Roach and John D. McNeish1 -- 2 The Use of a Chemically Defined Media for the Analyses of Early -- Development in ES Cells and Mouse Embryos -- Gabriele Proetzel and Michael V. Wiles17 -- 3 Analysis of the Cell Cycle in Mouse Embryonic Stem Cells -- Pierre Savatier, H6elne Lapillonne, Ludmila Jirmanova, -- Luigi Vitelli, and Jacques Samarut27 -- 4 Murine Embryonic Stem Cells as a Model for Stress Proteins -- and Apoptosis During Differentiation -- Andre-Patrick Arrigo and Patrick Mehlen35 -- 5 Effects of Altered Gene Expression on ES Cell Differentiation -- Yong Fan and J. Richard Chaillet45 -- 6 Hypoxic Gene Regulation in Differentiating ES Cells -- David M. Adelman and M. Celeste Simon55 -- 7 Regulation of Gap Junction Protein (Connexin) Genes and Function -- in Differentiating ES Cells -- Masahito Oyamada, Yumiko Oyamada, Tomoyuki Kaneko, -- and Tetsuro Takam atsu63 -- 8 Embryonic Stem Cell Differentiation as a Model to Study -- Hematopoietic and Endothelial Cell Development -- Stuart T. Fraser, Minetaro Ogawa, Satomi Nishikawa, -- and Shin-lchi N ishikaw a71 -- 9 Analysis of Bcr-AblFunction Using an In Vitro Embryonic Stem Cell -- Differentiation System -- Takumi Era, Stephane Wong, and Owen N. Witte 83 -- 10 Embryonic Stem Cells as a Model for Studying Osteoclast Lineage -- Development -- Toshiyuki Yamane, Takahiro Kunisada, and Shin-lchi Hayashi 97 -- 11 Differentiation of Embryonic Stem Cells as a Model to Study Gene -- Function During the Development of Adipose Cells -- C hristian D ani107 -- 12 Embryonic Stem Cell Differentiation and the Vascular Lineage -- V ictoria L. B autch117 -- 13 Embryonic Stem Cells as a Model to Study Cardiac, -- Skeletal Muscle, and Vascular Smooth Muscle Cell Differentiatiin -- Anna M. Wobus, Kaomei Guan, Huang-Tian Yang, -- and Kenneth R. Boheler127 -- 14 Cardiomyocyte Enrichment in Differentiating ES Cell Cultures: -- Strategies and Applications -- Kishore B. S. Pasumarthi and Loren J. Field157 -- 15 Embryonic Stem Cells as a Model for the Physiological Analysis -- of the Cardiovascular System -- Jurgen Hescheler, Maria Wartenberg, Bernd K. Fleischmann, -- Kathrin Banach, Helmut Acker, and Heinrich Sauer 169 -- 16 Isolation of Lineage-Restricted Neural Precursors from Cultured -- ES Cells -- Tahmina Mujtaba and Mahendra S. Rao189 -- 17 Lineage Selection for Generation and Amplification of Neural -- Precursor Cells -- M eng L i2 0 5 -- 18 Selective Neural Induction from ES Cells by Stromal -- Cell-Derived InducingActivity and Its Potential Therapeutic -- Application in Parkinson's Disease -- Hiroshi Kawasaki, Kenji Mizuseki, and Yoshiki Sasai217 -- 19 Epidermal Lineage -- Tammy-Claire Troy and Kursad Turksen229 -- 20 ES Cell Differentiation Into the Hair Follicle Lineage In Vitro -- Tammy-Claire Troy and Kursad Turksen255 -- 21 Embryonic Stem Cells as a Model for Studying Melanocyte -- Development -- Toshiyuki Yamane, Shin-lchi Hayashi, and Takahiro Kunisada 261 -- 22 Using Progenitor Cells and Gene Chips to Define Genetic Pathways -- S. Steven Potter, M. Todd Valerius, and Eric W. Brunskill 269 -- 23 ES Cell-Mediated Conditional Transgenesis -- Marina Gertsenstein, Corrinne Lobe and Andras Nagy 285 -- 24 Switching on Lineage Tracers Using Site-Specific Recombination -- Susan M. Dymecki, Carolyn I. Rodriguez, -- and Rajeshwar B. Awatramani309 -- 25 From ES Cells to Mice: The Gene Trap Approach -- Francesco Cecconi and Peter Gruss 335 -- 26 Functional Genomics by Gene-Trapping in Embryonic Stem Cells -- Thomas Floss and Wolfgang Wurst347 -- 27 Phage-Displayed Antibodies to Detect Cell Markers -- Jun Lu and Steven R. Sloan 381 -- 28 Gene Transfer Using Targeted Filamentous Bacteriophage if -- David Larocca, Kristen Jensen-Pergakes, Michael A. Burg, -- and A ndrew Baird 393 -- 29 Single-Cell PCR Methods for Studying Stem Cells and Progenitors -- Jane E. Aubin, Fina Liu, and G. Antonio Candeliere 403 -- 30 Nonradioactive Labeling and Detection of mRNAs Hybridized -- onto Nucleic Acid cDNA Arrays -- Thorsten Hoevel and Manfred Kubbies417 -- 31 Expression Profiling Using Quantitative Hybridization -- on Macroarrays -- Genevieve Pietu and Charles Decraene 425 -- 32 Isolation of Antigen-Specific Intracellular Antibody Fragments -- as Single Chain Fv for Use in Mammalian Cells -- Eric Tse, Grace Chung, and Terence H. Rabbitts433 -- 33 Detection and Visualization of Protein Interactions with Protein -- Fragment Complementation Assays -- Ingrid Remy, Andre Galarneau, and Stephen W. Michnick447 -- 34 Direct Selection of cDNAs by Phage Display -- Reto Crameri, Gernot Achatz, Michael Weichel, -- and Claudio Rhyner461 -- 35 Screening for Protein-Protein Interactions in the Yeast -- Two-Hybrid System -- R. Daniel Gietz and Robin A. Woods471 -- Index 4 87 Methods For The Isolation And Maintenance Of Murine Embryonic Stem Cells / Marsha L. Roach -- The Use Of A Chemically Defined Media For The Analyses Of Early Development In Es Cells And Mouse Embryos / Gabriele Proetzel -- Analysis Of The Cell Cycle In Mouse Embryonic Stem Cells / Pierre Savatier -- Murine Embryonic Stem Cells As A Model For Stress Proteins And Apoptosis During Differentiation / André-patrick Arrigo -- Effects Of Altered Gene Expression On Es Cell Differentiation / Yong Fan -- Hypoxic Gene Regulation In Differentiating Es Cells / David M. Adelman -- Regulation Of Gap Junction Protein (connexin) Genes And Function In Differentiating Es Cells / Masahito Oyamada -- Embryonic Stem Cell Differentiation As A Model To Study Hematopoietic And Endothelial Cell Development / Stuart T. Fraser -- Analysis Of Bcr-abl Function Using An In Vitro Embryonic Stem Cell Differentiation System / Takumi Era --^ Embryonic Stem Cells As A Model For Studying Osteoclast Lineage Development / Toshiyuki Yamane -- Differentiation Of Embryonic Stem Cells As A Model To Study Gene Function During The Development Of Adipose Cells / Christian Dani -- Embryonic Stem Cell Differentiation And The Vascular Lineage / Victoria L. Bautch -- Embryonic Stem Cells As A Model To Study Cardiac, Skeletal Muscle, And Vascular Smooth Muscle Cell Differentiation / Anna M. Wobus -- Cardiomyocyte Enrichment In Differentiating Es Cell Cultures: Strategies And Applications / Kishore B.s. Pasumarthi -- Embryonic Stem Cells As A Model For The Physiological Analysis Of The Cardiovascular System / Jürgen Hescheler -- Isolation Of Lineage-restricted Neural Precursors From Cultured Es Cells / Tahmina Mujtaba -- Lineage Selection For Generation And Amplification Of Neural Precursor Cells / Meng Li --^ Selective Neural Induction From Es Cells By Stromal Cell-derived Inducing Activity And Its Potential Therapeutic Application In Parkinson's Disease / Hiroshi Kawasaki -- Epidermal Lineage / Tammy-claire Troy -- Es Cell Differentiation Into The Hair Follicale Lineage In Vitro / Tammy-claire Troy -- Embryonic Stem Cells As A Model For Studying Melanocyte Development / Toshiyuki Yamane -- Using Progenitor Cells And Gene Chips To Define Genetic Pathways / S. Steven Potter -- Es Cell-mediated Conditional Transgenesis / Marina Gertsenstein -- Switching On Lineage Tracers Using Site-specific Recombination / Susan M. Dymecki -- From Es Cells To Mice: The Gene Trap Approach / Francesco Cecconi -- Functional Genomics By Gene-trapping In Embryonic Stem Cells / Thomas Floss -- Phage-displayed Antibodies To Detect Cell Markers / Jun Lu -- Gene Transfer Using Targeted Filamentous Bacteriophage / David Larocca -- Single-cell Pcr Methods For Studying Stem Cells And Progenitors / Jane E. Aubin --^ Nonradioactive Labeling And Detection Of Mrnas Hybridized Onto Nucleic Acid Cdna Arrays / Thorsten Hoevel -- Expression Profiling Using Quantitative Hybridization On Macroarrays / Geneviéve Piétu -- Isolation Of Antigen-specific Intracellular Antibody Fragments As Single Chain Fv For Use In Mammalian Cells / Eric Tse -- Detection And Visualization Of Protein Interactions With Protein Fragment Complementation Assays / Ingrid Remy -- Direct Selection Of Cdnas By Phage Display / Reto Crameri -- Screening For Protein-protein Interactions In The Yeast Two-hybrid System / R. Daniel Gietz. Edited By Kursad Turksen. Includes Bibliographical References And Index. Kursad Turksen and a panel of international experts describe their most productive methods for using embryonic stem (ES) cells as in vitro developmental models for many cell and tissue types. Set out in step-by-step detail, these protocols range widely from ES cell isolation, maintenance, and modulation of gene expression, to cutting-edge techniques that use cDNA arrays in gene expression analysis and phage display libraries, the generation of antibodies against very rare antigens, and the identification of and characterization of protein and protein interactions. Embryonic Stem Cells: Methods and Protocols will prove an invaluable resource not only for those interested generally in cell and developmental biology, but also for those actively using, or planning to use ES cells, to study fate choices and specific lineages. Annotation Kursad Turksen and a panel of international experts describe their most productive methods for using embryonic stem (ES) cells as in vitro developmental models for many cell and tissue types. Set out in step-by-step detail, these protocols range widely from ES cell isolation, maintenance, and modulation of gene expression, to cutting-edge techniques that use cDNA arrays in gene expression analysis and phage display libraries, the generation of antibodies against very rare antigens, and the identification of and characterization of protein and protein interactions