Optimization of a large-scale gene disruption protocol in Dictyostelium and analysis of conserved genes of unknown function.

ABSTRACT:

Background

Development of the post-genomic age in Dictyostelium will require the existence of rapid and reliable methods to disrupt genes that would allow the analysis of entire gene families and perhaps the possibility to undertake the complete knock-out analysis of all the protein-coding genes present in Dictyostelium genome.

Results

Here we present an optimized protocol based on the previously described construction of gene disruption vectors by in vitro transposition. Our method allows a rapid selection of the construct by a simple PCR approach and subsequent sequencing. Disruption constructs were amplified by PCR and the products were directly transformed in Dictyostelium cells. The selection of homologous recombination events was also performed by PCR. We have construc

SUBMITTER: Torija P 

PROVIDER: S-EPMC1564143 | biostudies-literature | 2006 Aug

REPOSITORIES: biostudies-literature