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Title: A non-destructive seed sampling method for PCR-based analyses in marker assisted selection and transgene screening

Author
item Chamberlin, Kelly
item GALLO, MARIA - UNIV OF FLORIDA
item SEIB, JEFFERY - UNIV OF FLORIDA
item JAMES, V - UNIV OF FLORIDA

Submitted to: Peanut Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/1/2006
Publication Date: 6/1/2007
Citation: Chenault, K.D., Gallo, M., Seib, J.C., James, V.A. 2007. A non-destructive seed sampling method for PCR-based analyses in marker assisted selection and transgene screening. Peanut Science. 34:38-43.

Interpretive Summary: Extraction of DNA from peanut tissue generally requires extensive manipulation in order to remove numerous compounds which interfere with molecular techniques. To reduce the amount of these compounds present, routine purification of peanut DNA is normally performed on young leaf tissue which requires time consuming seed germination and growth of the peanut plant for at least two to four weeks. We have developed a quick method to extract DNA directly from a small portion of the peanut seed, leaving the seed viable for germination and further use. This method provides enough quality DNA to perform molecular analysis. Since this method is non-destructive, seed can be subsequently germinated to produce healthy, mature plants, making this technique a useful tool in screening populations of peanut seed for desirable traits that can be used to enhance breeding programs.

Technical Abstract: Extraction of quality DNA from peanut (Arachis hypogaea L.) generally requires extensive manipulation in order to remove numerous phenolic compounds and polysaccharides. To reduce the amount of problematic compounds present, routine purification of peanut DNA is normally performed on young leaf tissue which requires time consuming seed germination and growth of the peanut plant for at least two to four weeks. The extraction method described here uses 0.02 g of peanut cotyledon tissue taken directly from the distal end of the mature seed and provides 30-46 micro g of DNA suitable for use in restriction digests, PCR, SCAR and SSR analyses. Since this method is non-destructive, seed can be subsequently germinated to produce healthy, mature plants, making this technique a useful tool for screening segregating populations of potentially transgenic seed and in breeding programs that use molecular marker assisted selection to screen for desired traits.