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Title: Modified Ecotilling protocol for simple and rapid SNP discovery and genotyping in rice

Author
item KADARU, SURESH - LSU AGCENTER
item YADAV, ANIRUDHA - DEPT. OF BOTANY - INDIA
item Fjellstrom, Robert
item OARD, JAMES - LSU AGCENTER

Submitted to: Plant and Animal Genome Conference Proceedings
Publication Type: Proceedings
Publication Acceptance Date: 12/1/2005
Publication Date: 1/1/2006
Citation: Kadaru, S.B., Yadav, A.S., Fjellstrom, R.G., Oard, J.H. 2006. Modified Ecotilling protocol for simple and rapid SNP discovery and genotyping in rice [abstract]. Plant and Animal Genome Conference Proceedings. p. P255.

Interpretive Summary:

Technical Abstract: Ecotilling is a high through-put method for detection and analysis of DNA variation in large populations, but requires a substantial investment in equipment and reagents for laboratories with limited financial resources. Modifications to Ecotilling were optimized in rice to address this issue using a simplified gel system, conventional PCR, and standard data analysis that resulted in rapid, cost efficient SNP discovery and genotyping. To validate the new procedure, four SNPs previously reported in the rice alk and waxy genes were detected and shown to be faithfully genotyped among 57 diverse rice accessions. Analysis based on direct sequencing of 30 rice accessions in the alk and waxy gene regions further confirmed accuracy of SNP analysis by the modified Ecotilling approach. Associations between SNPs and amylose class starch gelatinization temperature were as expected. Detection of a single SNP among a large diverse group of rice accessions suggested that modified Ecotilling could identify polymorphisms at low frequencies in natural populations. This technique also validated an insertion/deletion in the first intron of the waxy gene. A detailed time and cost analysis showed that modified Ecotilling is a quick, simple, and inexpensive protocol for SNP discovery and genotyping in natural selected populations of rice.