Nature Biotechnology23, 75 - 81 (2004)
Published online: 5 December 2004; | doi:10.1038/nbt1043
A reverse genetic, nontransgenic approach to wheat crop improvement by TILLING
Ann J Slade1, Susan I Fuerstenberg2, Dayna Loeffler1, Michael N Steine1
& Daniel Facciotti2
1
Anawah Inc., 1102 Columbia Street, Suite 600, Seattle Washington 98104, USA.
2
Anawah Inc., 215 Madson Place, Davis, California 95616, USA.
Correspondence should be addressed to Ann J Slade ann@anawah.com
We report the use of TILLING (targeting induced local lesions in genomes), a reverse genetic, nontransgenic method, to improve a quality trait in a polyploid crop plant. Waxy starches, composed mostly of amylopectin, have unique physiochemical properties. Wheat with only one or two functional waxy genes (granule-bound starch synthase I, or GBSSI) produces starch with intermediate levels of amylopectin. We have identified 246 alleles of the waxy genes by TILLING each homoeolog in 1,920 allohexaploid and allotetraploid wheat individuals. These alleles encode waxy enzymes ranging in activity from near wild type to null, and they represent more genetic diversity than had been described in the preceding 25 years. A line of bread wheat containing homozygous mutations in two waxy homoeologs created through TILLING and a preexisting deletion of the third waxy homoeolog displays a near-null waxy phenotype. This approach to creating and identifying genetic variation shows potential as a tool for crop improvement.
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