ArticleNucleic acids research2003
Rapid generation of gene disruption constructs by in vitro transposition and identification of a Dictyostelium protein kinase that regulates its rate of growth and development.
Article in Nucleic acids research, 2003. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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23 citing papers in PubMed, 35 citations in OpenAlex.
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- ABC transporters in Dictyostelium discoideum development.PloS one · 2013Article
- A secreted protein is an endogenous chemorepellant in Dictyostelium discoideum.Proceedings of the National Academy of Sciences of the United States of America · 2012Article
- The LRRK2-related Roco kinase Roco2 is regulated by Rab1A and controls the actin cytoskeleton.Molecular biology of the cell · 2011Article
- Cell type-specific filamin complex regulation by a novel class of HECT ubiquitin ligase is required for normal cell motility and patterning.Development (Cambridge, England) · 2011Article
- The ROCO kinase QkgA is necessary for proliferation inhibition by autocrine signals in Dictyostelium discoideum.Eukaryotic cell · 2010Article
- Two novel Src homology 2 domain proteins interact to regulate dictyostelium gene expression during growth and early development.The Journal of biological chemistry · 2010Article
- Characterization of the Roco protein family in Dictyostelium discoideum.Eukaryotic cell · 2010Article
- Regulation of Rap1 activity is required for differential adhesion, cell-type patterning and morphogenesis in Dictyostelium.Journal of cell science · 2009Article
- Vacuole membrane protein 1 is an endoplasmic reticulum protein required for organelle biogenesis, protein secretion, and development.Molecular biology of the cell · 2008Article
- DIF-1 induces the basal disc of the Dictyostelium fruiting body.Developmental biology · 2008Article
- Proteomic and microarray analyses of the Dictyostelium Zak1-GSK-3 signaling pathway reveal a role in early development.Eukaryotic cell · 2007Article
- Biosynthesis of Dictyostelium discoideum differentiation-inducing factor by a hybrid type I fatty acid-type III polyketide synthase.Nature chemical biology · 2006Article
- Optimization of a large-scale gene disruption protocol in Dictyostelium and analysis of conserved genes of unknown function.BMC microbiology · 2006Article
- The dictyostelium kinome--analysis of the protein kinases from a simple model organism.PLoS genetics · 2006Article
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3 authors at 1 institution in 1 country.
Funding
Abstract
We describe a rapid method for creating Dictyo stelium gene disruption constructs, whereby the target gene is interrupted by a drug resistance cassette using in vitro transposition. A fragment of genomic DNA containing the gene to be disrupted is amplified by PCR, cloned into a plasmid vector using topoisomerase and then employed as the substrate in an in vitro Tn5 transposition reaction. The transposing species is a fragment of DNA containing a Dictyostelium blasticidin S resistance (bs(r)) cassette linked to a bacterial tetracycline resistance (tet(r)) cassette. After transposition the plasmid DNA is transformed into Escherichia coli and clones in which the bs(r)-tet(r) cassette is inserted into the Dictyostelium target DNA are identified. To demonstrate its utility we have employed the method to disrupt the gene encoding QkgA, a novel protein kinase identified from the Dictyostelium genome sequencing project. QkgA is structurally homologous to two previously identified Dictyostelium kinases, GbpC and pats1. Like them it contains a leucine-rich repeat domain, a small GTP-binding (ras) domain and a MEKK domain. Disruption of the qkgA gene causes a marked increase in growth rate and, during development, aggregation occurs relatively slowly to form abnormally large multicellular structures.
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