In Drosophila Cytogenetics Protocols, leading drosophilists describe, in step-by-step detail, all the essential techniques for studying Drosophila chromosomes and suggest new avenues for scientific exploration.
General areas covered include analyses of nuclei and chromosomes in spermatogenesis and oogenesis, fluorescence-activated cell sorting of ovarian follicle cells, analyses of nuclear division cycles in embryos, and analyses of programmed cell death. In addition, there are conventional light-, electron-, and immunofluorescence-microscopic analyses of polytene chromosomes; cytological analyses of larval brains, imaginal discs, and histoblasts; and nonfluorescent and fluorescent in situ hybridization FISH to both polytene and mitotic chromosomes.
A cutting-edge combined FISH and immunolocalization protocol will be important for discovering how DNA sequence influences higher-order chromosome architecture and ultimately gene expression. Each readily reproducible protocol offers a background introduction, equipment and reagent lists, and tips on troubleshooting and avoiding pitfalls. Readily accessible and highly practical, Drosophila Cytogenetics Protocols provides a comprehensive cytogenetics laboratory manual for Drosophila investigators, one that is suitable not only for novices, but also highly informative for seasoned investigators.
Leading drosophilists describe in step-by-step detail all the essential techniques for studying Drosophila chromosomes and suggest new avenues for scientific exploration. Each fully tested and readily reproducible protocol offers a background introduction, equipment and reagent lists, and tips on troubleshooting and avoiding pitfalls. A cutting-edge FISH and immunolocalization technique will be important for discovering how DNA sequence influences higher-order chromosome architecture and ultimately gene expression.
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The normal development of an egg chamber has been divided into 14 stages 4. Stages are "previtellogenic," with intensive growth of all the cells that form each chamber. At stage 7, the process of transfer of yolk precursors from NCs into the oocyte begins.
Vitellogenesis, accelerated growth by the oocyte. From: Methods in Molecular Biology, vol.
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By stage 9, NCs also develop unusual nucleoli composed of a shell of interconnected fibers around the periphery of the nucleus. Stage 11 is the shortest and marks the beginning of the postvitellogenic period, with apoptosis of NCs, and the completion of the vitelline membrane.
Drosophila Cytogenetics Protocols
During stages , beta yolk forms, and the egg shell and its appendages are synthesized. The staging characteristics for Drosophila oogenesis were formulated after observations of Feulgen-stained, ovarian whole mounts 8 see Subheading 3. The NC nuclei in chambers belonging to stages possessed a dispersed mass of Feulgen-positive threads see Fig. Nuclei in stages 3 and 4 contained densely staining bulbous structures see Fig.
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The NCs in stage 5 were at an obviously intermediate state. The nuclei nearest the oocyte contained dispersed chromosomes like those in stage 6, whereas the rest resembled stage 4. At stage 5, the bivalents fall apart, starting with the NCs closest to the oocyte, and the polytene univalents subdivide further to form 32 separate chromatids, each containing the 2C amount of DNA and held together by unreplicated regions.
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There are three to four replication cycles, depending on the distance of the NC from the oocyte.