Chapter 17 Cytoplasmic Extracts from the Eggs of Sea Urchins and Clams for the Study of Microtubule-Associated Motility and Bundling

Abstract

Publisher Summary This chapter describes clarified cytoplasmic extracts made from the eggs of sea urchins or clams that are used to study microtubule-associated translocation and bundling in the presence of soluble cytoplasmic proteins. In the extracts, microtubules made from endogenous tubulin both translocate across coverslips and bundle to each other. In addition, the microtubules serve as a substrate for the bidirectional movement of organelles. Sea urchin oocytes and eggs are a rich source of the cytoskeletal components necessary for the rapid cell divisions following fertilization. Three of the materials stockpiled for postfertilization events include tubulin, microtubule-associated proteins, and microtubule motor proteins. Sea urchin egg fertilization should be carried out in ATA seawater to allow for removal of the fertilization membrane. The oocytes of the surf clam Spisula solidissima can be collected in quantities suitable for biochemical studies and are excellent cytological specimens. The mature Spisula oocyte has an intact germinal vesicle with a tetraploid complement of partially condensed chromosomes. The proximity in time between fertilization and the first meiotic division allows the harvesting of cells in high cell cycle synchrony.