Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
2821069 | Genomics | 2011 | 5 Pages |
The amoeba Dictyostelium discoideum is a well-established model organism for studying numerous aspects of cellular and developmental functions. Its rather small (~ 34 Mb) chromosomal genome and the high efficiency of gene disruption by homologous recombination have enabled researchers to dissect various specific gene functions. We describe here the use of one-step cloning for the fast and efficient generation of deletion vectors that are produced in a one-step reaction by inserting two PCR products into an organism-specific, generic acceptor system. This worked efficiently for all 16 tested constructs directed against genes in the amoeba Dictyostelium discoideum. Saving cost and time, the used protocol represents a significant advancement in the generation of such plasmids compared to the conventionally applied restriction enzyme/ligation approach. Using appropriate selection markers, similar systems could also be useful in other organisms, where genes can be knocked out by homologous recombination.