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CELL BIOLOGY & MOLECULAR GENETICS

Modified Rapid DNA Extraction Protocol for High Throughput Microsatellite Analysis in Wheat

USDA-ARS Plant Science and Entomology Unit, 4008 Throckmorton Hall, Manhattan, KS 66506

* Corresponding author (jpf{at}alfalfa.ksu.edu)

New technology is allowing marker-assisted selection to fulfill the promise of increasing efficiency of cultivar development. However, these techniques depend upon the ability to extract DNA from large populations of plants. The objective of this project was to develop a high-throughput DNA extraction procedure without the need for greenhouse space or growing wheat (Triticum aestivum L.) plants. A sodium hydroxide rapid DNA extraction was modified for a 96-well format to reduce costs. Seeds were germinated in 8-well tissue culture plates, and 4-d-old seedling tissue was used to extract DNA by means of sodium hydroxide methodology. Approximately 1 µg of genomic DNA per 10 mg of tissue was isolated at a cost of about $0.10. The DNA quality was verified by amplification of microsatellite markers. Results were consistent with either fresh or stored tissue extracts. This technique allows one person to extract nearly 1000 storage-stable DNA samples daily, while keeping costs at a minimum.

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