Isolation of Nuclei From Xenopus Oocytes

A piece of ovary containing mature (stage VI) oocytes is surgically removed from female X. laevis and stored in MBS at 4°C for a maximum of 5 d. Before isolation of the nuclei, individual oocytes are placed (with 1-mL pipet with cut off tip's end; see Note 3) in a small Petri dish containing LSB to equilibrate them at room temperature. This buffer is commonly used to keep nuclei intact and partially swollen (nuclei are easily popped out) during the isolation. Nuclei are manually isolated either by opening the oocytes at their animal (dark) pole or by disrupting the oocytes with fine Dumont forceps along the line separating the two hemispheres. The released nuclei are cleaned from yolk and other materials by sucking them gently up and down into the tip of a 20-|L pipet (adjusted to 5 ||L; see Note 3) in LSB containing a trace amount of PMSF (see Note 1) (5,8).

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