Int. J. Dev. Biol. 46: 309 - 316 (2002)

Vol 46, Issue 3

Selecting karyophilic DNA cis elements in Xenopus laevis oocytes; a new approach

Published: 1 May 2002

Marco Marcello, Roger Fischer, Helmut Tröster, Michael F Trendelenburg and Georg Sczakiel

Deutsches Krebsforschungszentrum, Heidelberg, Germany.

Abstract

The intracellular localisation and mobility of exogenous DNA introduced into Xenopus laevis oocytes is largely unknown. In this paper, we report a new technique to investigate the cytoplasmic/nuclear transport of a random pool of linear, double-stranded, oligomeric DNA of 147 bp in length. We chose a combinatorial approach which made use of repetitive rounds of selection and amplification to search for new cis elements mediating nuclear import or retention. A new PCR-based methodology was established to reliably detect exogenous DNA in subcellular and total extracts prepared from Xenopus laevis oocytes. Studies in vivo and with cellular extracts indicate the presence of a highly efficient nuclease activity in the nuclear compartment. The described combinatorial approach constitutes a promising tool for the isolation of novel DNA cis elements which may play an important role in the nuclear internalisation and retention of exogenous DNA in Xenopus laevis oocytes.