Michael Gore1, Peter Bradbury 2, René Hogers 3, Matias Kirst 4, Esther Verstege3, Jan van Oeveren3, Johan Peleman3, Edward Buckler 5, and Michiel J.T. van Eijk3. (1) Institute for Genomic Diversity , Cornell University , 175 Biotechnology Building , Ithaca , NY 14853-2703 , USA , (2) USDA-ARS , Cornell University , 741 Rhodes Hall , Ithaca , NY 14853 , USA , (3) Keygene N.V. , Agro Business Park 90 , P.O. Box 216 , 6700 AE Wageningen , , Netherlands , (4) School of Forest Resources and Conservation , University of Florida , PO Box 110410 , Gainesville , FL 32610 , USA , (5) USDA-ARS , Institute for Genomic Diversity , Cornell University , Ithaca , NY 14853-2703 , USA
We describe the detection of sequence validated single-feature polymorphisms (SFPs) between maize inbred lines by hybridizing RNA or complexity-reduced genomic DNA to an Affymetrix GeneChip® expression array. Direct hybridization of labeled total genomic DNA to oligonucleotide expression arrays for SFP detection was initially demonstrated in organisms with relatively small genomes, such as ~12Mb yeast (Winzeler et al., 1998) and 130Mb Arabidopsis (Borevitz et al., 2003). For larger genomes such as ~2500Mb maize, however, obtaining robust hybridization signals requires a sample preparation method optimized to reduce repetitive or high-copy sequences in the target prior to array hybridization. In this study, we evaluated the efficacy of four different complexity-reduction methods for sensitive SFP detection in maize: cDNA, methyl filtration, high-Cot selection, and AFLP®. These four methods were applied to four diverse maize inbred lines (B73, Mo17, CML69 and Tzi8) with 3 replications per line (48 GeneChips®). Our results indicate that all genome reduction methodologies offer modest power to detect SFPs with the commercially available GeneChip® Maize Genome Array. Moreover, the detection of non-SNP genetic polymorphisms by probes, such as allelic non-homologies and copy number of genes, may produce some of the apparent Type I error rates.
The AFLP technology is covered by patents and patent applications owned by Keygene. AFLP® is a registered trademark of Keygene.
Back to Graduate Student Paper Contest (Oral)
Back to The Northeastern Branch of the ASA-SSSA (July 11, 2006)
