Blasticidin resistance cassette in symmetrical polylinkers for insertional inactivation of genes in Dictyostelium.
In Dictyostelium discoideum inactivation of developmentally regulated genes via homologous recombination has become an important tool in studying systematically the entire developmental program of this model organism. The Dictyostelium genome is very A/T-rich,which presents obstacles to the preparation of knockout constructs. The coding regions offer few suitable restriction sites and the low complexity intergenic regions do not guarantee specificity of recombination. We present here the preparation of plasmids pBsR479, pBsR503, and pBsR519, in which a blasticidin resistance-cassette is positioned in the center of various symmetrical polylinkers. This design simplifies the cloning process and gives more flexibility in positioning the selectable marker within the coding regions.
Půta
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F
Department of Physiology and Developmental Biology, Faculty of Science, Charles University, Prague, Czech Republic.
Zeng
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C
D000818:Animals; D001483:Base Sequence; D016054:DNA, Protozoan / Q000235:genetics; D004023:Dictyostelium / Q000187:drug effects* / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics; D018390:Gene Targeting / Q000379:methods*; D017125:Genes, Protozoan; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000379:methods*; D009705:Nucleosides / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D015183:Restriction Mapping
D016054:DNA, Protozoan; D009705:Nucleosides; C004500:blasticidin S