A cell-free system of Drosophila egg explants supporting native mitotic cycles

• Published in: Methods in cell biology Vol. 144; pp. 233 - 257
• Main Authors: de-Carvalho, Jorge, Deshpande, Ojas, Nabais, Catarina, Telley, Ivo A
• Format: Book Chapter Journal Article
• Language: English
• Published: United States Elsevier Science & Technology 2018
• Subjects: Animals; Cell Extracts; Cell-Free System; Cytological Techniques - methods; Drosophila melanogaster - cytology; Drosophila melanogaster - genetics; Female; Image Processing, Computer-Assisted; Microscopy; Mitosis; Ovum - cytology; Time Factors; Mitotic cycling; Cell-free; Extract; Ex vivo; Nuclear division; Drosophila
• ISBN: 9780128141441; 0128141441
• ISSN: 0091-679X
• DOI: 10.1016/bs.mcb.2018.03.011

Mitosis, in a broader sense, is an intracellular mechanical process that is fueled by chemical reactions and regulated by a complex protein interaction network. Research aimed at understanding mitosis in all these aspects is often limited to pharmaceutical treatment or genetic manipulation of single cells or entire tissues. These experimental models entail physical boundaries imposed by the cell membrane, making it extremely challenging to apply mechanical perturbations, or to introduce larger molecules such as peptides, proteins, or genetic transcripts in an acute and specific manner. Here, we present a cell-free experimental assay that is exploiting the properties of a large, multinucleated embryo cell. Drosophila, like almost all insects, initially develops as a syncytial embryo, the task of which is to replicate and distribute the genetic material quickly and regularly. We describe an experimental procedure that allows the isolation of nucleocytoplasm from single embryos that retains the developmental processes, most importantly the native mitotic progression of nuclei.