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Genetic Engineering of Dictyostelium discoideum Cells Based on Selection and Growth on Bacteria
Published on: January 25, 2019
A reliable general purpose method for extracting genomic DNA from Dictyostelium cells
Karen E Pilcher1, Petra Fey, Pascale Gaudet
1dictyBase, Center for Genetic Medicine, Northwestern University, 676 North Saint Clair Street Suite 1260, Chicago, Illinois 60611, USA.
Nature Protocols
|June 5, 2007
Summary
This protocol details a 3-hour DNA extraction method for Dictyostelium discoideum cells. It effectively isolates genomic DNA from both wild-type and modified strains for molecular analyses.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Dictyostelium discoideum presents unique challenges for DNA extraction due to high carbohydrate and nuclease content.
- Standard phenol:chloroform methods require modification for efficient DNA isolation from this organism.
Purpose of the Study:
- To present a standardized, efficient protocol for genomic DNA extraction from Dictyostelium discoideum.
- To enable molecular genetic analyses in both wild-type and genetically modified Dictyostelium strains.
Main Methods:
- The protocol involves cell lysis, nucleic acid extraction, and precipitation.
- Modifications are incorporated to overcome challenges posed by high carbohydrate and nuclease levels in Dictyostelium.
Main Results:
- The method successfully isolates genomic DNA suitable for downstream applications.
- Isolated DNA is compatible with restriction enzyme digestion, PCR amplification, and Southern blotting.
Conclusions:
- This optimized protocol provides a reliable method for DNA extraction in Dictyostelium.
- The procedure facilitates various molecular genetic studies using Dictyostelium discoideum.
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