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United States Department of Agriculture

Agricultural Research Service

Title: Discovery of Genomic DNA Polymorphisms using oligonucleotide arrays

Authors
item Leong, Sally
item Tian, S - UNIV OF WISCONSIN
item Splinter-Bondurant, S - UNIV OF WISCONSIN

Submitted to: Rice Technical Working Group Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: February 26, 2006
Publication Date: February 26, 2006
Citation: Leong, S.A., Tian, S., Splinter-Bondurant, S. 2006. Discovery of Genomic DNA Polymorphisms using oligonucleotide arrays. Rice Technical Working Group Meeting Proceedings. p. 56.

Technical Abstract: We are applying oligonucleotide microarray technology as a means of rapidly discovering DNA-based markers for genetic mapping in finger millet and rice. A test genome scan was conducted using the full genome sequences of rice varieties 93-11 (indica) and Nipponbare (japonica) compared with the Affymetrix rice gene array probe set to determine which probes are polymorphic. A total of 93,353 putative single feature polymorphisms were identified in the 93-11 genome based on our genome scanning definitions, indicating that this approach should be fruitful. A random priming method for labeling of the genomic DNA target was optimized and test hybridizations were conducted with labeled genomes of the two rice genotypes 93-11 and Nipponbare. Analysis of this preliminary, un-replicated data using several data normalization methods showed that a large percentage of the putative SFPs can be detected when single copy probe data are considered. For example, using a signal cutoff of equal to or greater than 200 for the probes from the reference genome, as well as a ratio of 2 or greater for the perfect match signal for the Nipponbare target over the perfect match signal for the 93-11 target, 11,019 single copy SFPs were found based on these criteria. Of these 8,676 do not have a 25 bp perfect match in the 93-11 genome based on genome scanning, yielding a specificity of 78.7%. Using a ratio of 3 or greater, 4,925 putative SFPs were identified, of which 4,177 were identified through genome scanning, yielding a specificity of 84.8%. Increasing signal to noise and insuring probe saturation are possible improvements to strive for in future. Application of this technology to the discovery of rice single feature polymorphisms that can be used for genetic mapping of quantitative resistance to sheath blight and milling quality and yield will be discussed.

Last Modified: 7/28/2014
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