Methodology article

Detection of genomic deletions in rice using oligonucleotide microarrays

Myron Bruce¹ , Ann Hess² , Jianfa Bai³ , Ramil Mauleon⁴ , M Genaleen Diaz^1,5 , Nobuko Sugiyama⁴ , Alicia Bordeos⁴ , Guo-Liang Wang⁶ , Hei Leung⁴ and Jan E Leach¹

¹Program in Plant Molecular Biology, Department of Bioagricultural Sciences and Pest Management, Colorado State University, Fort Collins, USA

²Department of Statistics, Colorado State University, Fort Collins, USA

³Gene Expression Facility, Kansas State University, Manhattan, USA

⁴Internation Rice Research Institute, Manila, Philippines

⁵Philippines, Institute of Biological Sciences, University of the Philippines, Los Baños, Philippines

⁶Department of Plant Pathology, The Ohio State University, Columbus, USA

author email corresponding author email

BMC Genomics 2009, 10:129doi:10.1186/1471-2164-10-129

Published:	25 March 2009

Abstract

Background

The induction of genomic deletions by physical- or chemical- agents is an easy and inexpensive means to generate a genome-saturating collection of mutations. Different mutagens can be selected to ensure a mutant collection with a range of deletion sizes. This would allow identification of mutations in single genes or, alternatively, a deleted group of genes that might collectively govern a trait (e.g., quantitative trait loci, QTL). However, deletion mutants have not been widely used in functional genomics, because the mutated genes are not tagged and therefore, difficult to identify. Here, we present a microarray-based approach to identify deleted genomic regions in rice mutants selected from a large collection generated by gamma ray or fast neutron treatment. Our study focuses not only on the utility of this method for forward genetics, but also its potential as a reverse genetics tool through accumulation of hybridization data for a collection of deletion mutants harboring multiple genetic lesions.

Results

We demonstrate that hybridization of labeled genomic DNA directly onto the Affymetrix Rice GeneChip^® allows rapid localization of deleted regions in rice mutants. Deletions ranged in size from one gene model to ~500 kb and were predicted on all 12 rice chromosomes. The utility of the technique as a tool in forward genetics was demonstrated in combination with an allelic series of mutants to rapidly narrow the genomic region, and eventually identify a candidate gene responsible for a lesion mimic phenotype. Finally, the positions of mutations in 14 mutants were aligned onto the rice pseudomolecules in a user-friendly genome browser to allow for rapid identification of untagged mutations http://irfgc.irri.org/cgi-bin/gbrowse/IR64_deletion_mutants/ webcite.

Conclusion

We demonstrate the utility of oligonucleotide arrays to discover deleted genes in rice. The density and distribution of deletions suggests the feasibility of a database saturated with deletions across the rice genome. This community resource can continue to grow with further hybridizations, allowing researchers to quickly identify mutants that harbor deletions in candidate genomic regions, for example, regions containing QTL of interest.