Submitted to: Theoretical and Applied Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/4/2000
Publication Date: N/A
Citation: N/A Interpretive Summary: U.S. wheat and barley producers are facing serious problems in the quality of the grain they produce as a result of infection with Fusarium head blight (FHB) which also results in yield losses and may be associated with fungal toxins (mycotoxins) that are hazardous to animals. Resistance to FHB is controlled by a number of genes. Identifying and incorporating these genes into resistant varieties is a difficult task because of the difficulty of creating conditions under which resistance is reliably and consistently expressed. Molecular genetic markers provide an alternative to conventional disease resistance screening. Rather than selecting directly based upon the appearance of resistance, one selects for a form of the DNA marker that has been previously demonstrated to be closely associated with resistance. In order to permit this type of DNA marker aided selection, an informative set of DNA markers is needed. The objective of this work was to determine how to develop microsatellite DNA markers in wheat that would be maximally useful. Through the isolation and testing of nine different types of these markers we determined which types were frequent in the wheat genome and which could be used in the large scale development of informative microsatellite markers. This information is of immediate use to molecular biologists whose goal is to maximize the efficiency of microsatellite DNA marker development in wheat.
Technical Abstract: As compared to dinucleotide-based microsatellite markers, length differences among trinucleotide-based microsatellite alleles can be more easily detected and frequently produce fewer stutter bands. Our objective was to determine which trinucleotide motif(s) would be the most polymorphic and abundant source of trinucleotide microsatellite markers in wheat (Triticum aestivum L.). Four genomic libraries of cultivar Chinese Spring were screened with nine trinucleotide probes. Based on the screening of 28550 clones, the occurrences of (CTT/GAA)n, (GGA/CCT)n, (TAA/ATT)n, (CAA/GTT)n, (GGT/CCA)n, (CAT/GTA)n, (CGA/GCT)n, (CTA/GAT)n, and (CGT/GCA)n repeats were estimated to occur once every 293kbp, 456kbp, 500kbp, 1.3Mbp, 2.6Mbp, 3.2Mbp, 3.6Mbp, 3.6Mbp and 4.5Mbp in the wheat genome, respectively. Of 236 clones selected for sequencing, 38 (93%) (TAA/ATT)n, 30 (43%) (CTT/GAA)n, 16 (59%) (CAA/GTT)n, 3 (27%) (CAT/GTA)n and 2 (4%) (GGA/CCT)n clones contained microsatellites with 8 or more perfect repeats. From these data, 29, 27, and 16 PCR primer sets were designed and tested to the (TAA/ATT)n, (CTT/GAA)n, and (CAA/GTT)n microsatellites, respectively. A total of 12 (41.4%) primers designed to (TAA/ATT)n, 4 (14.8%) to (CTT/GAA)n, and 2 (12.5%) to (CAA/GTT)n resulted in polymorphic markers. The results indicated that (TAA/ATT)n microsatellites would provide the most abundant and the most polymorphic source of trinucleotide microsatellite markers in wheat.