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This detailed volume explores techniques for the isolation, purification, and establishment of in vitro germ cell (GS) systems at different stages of development in a variety of organisms, including chickens, mice, rats, and humans. The book additionally describes cutting-edge analytic and informatic tools to study germ cell development at the single cell level and meiotic recombination. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step and readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Authoritative and practical, Germ Cell Development: Methods and Protocols provides new insight into the understanding of basic biological aspects of germ cell biology, as well as the opportunity for in vitro manipulation of germ cells for toxicology studies and for genetic intervention.
List of contents
Isolation and Purification of Viable PGCs from Mouse Embryos.- Purification and Transfection Methods of Chicken Primordial Germ Cells.- Isolation and In Vitro Propagation of Human Spermatogonial Stem Cells.- Purification by STA-PUT Technique of Male Germ Cells from Single Mouse and RNA-Extraction for Transcriptomic Analysis.- Isolation of Mouse Germ Cells by FACS Using Hoechst 33342 and SYTO16 Double Staining.- Isolation and In Vitro Culture of Germ Cells and Sertoli Cells from Human Fetal Testis.- Human Primordial Germ Cell-Like Cell Induction from Pluripotent Stem Cells by SOX17 and PRDM1 Expression.- Induction of Primordial Germ Cell-Like cells from Rat Pluripotent Stem Cells.- Induction of Meiotic Initiation in Long-Term Mouse Spermatogonial Stem Cells Under Retinoid Acid and Nutrient Restriction Conditions.- Lipofection-Based Delivery of CRISPR/Cas9 Ribonucleoprotein for Gene Editing in Male Germline Stem Cells.- Mouse In Vitro Spermatogenesis on 3D Bioprinted Scaffolds.-Histological and Cytological Techniques to Study Perinatal Mouse Ovaries and Oocytes.- Method of Isolation and In Vitro Culture of Primordial Follicles in Bovine Animal Model.- Generation of iPSC Cell Lines from Patients with Sex Chromosome Aneuploidies.- Data Analysis Pipeline for scRNA-Seq Experiments to Study Early Oogenesis.- Mapping Meiotic DNA Breaks: Two Fully-Automated Pipelines to Analyze Single-Strand DNA Sequencing Data, hotSSDS and hotSSDSextra.- "MeiQuant": An Integrated Tool for Analyzing Meiotic Prophase I Spread Images.
