The density and utility of the molecular genetic linkage map of the widespread use of RFLP markers and maps in suncultivated sunflower (Helianthus annuus L.) has been greatly inflower has been restricted by a lack of public RFLP creased by the development and mapping of several hundred simple sequence repeat (SSR) markers. Of 1089 public SSR markers de-probes, consequent lack of a dense public RFLP map, scribed thus far, 408 have been mapped in a recombinant inbred line and low-throughput nature of RFLP markers. The diffi-(RIL) mapping population (RHA280 ϫ RHA801). The goal of the culties posed by the historic lack of public, single-copy present research was to increase the density of the sunflower map by DNA markers were only weakly offset by the emerconstructing a new RIL map (PHA ϫ PHB) based on SSRs, adding gence of facile, universal DNA markers, e.g., RAPDs loci for newly developed SSR markers to the RHA280 ϫ RHA801 RIL (Williams et al., 1990, 1993) and AFLPs (Vos et al., map, and integrating the restriction fragment length polymorphism 1995). RAPDs have primarily been used for tagging (RFLP) and SSR maps of sunflower. The latter was accomplished by phenotypic loci in sunflower, e.g., rust (Puccinia helianadding 120 SSR marker loci to a backbone of 80 RFLP marker loci thi Schw.) and Orobanche cumana Wallr. resistance on the HA370 ϫ HA372 F 2 map. The map spanned 1275.4 centimorgans (cM) and had a mean density of 6.3 cM per locus. The genes (Lawson et al., 1998; Lu et al., 2000). While RAPD PHA ϫ PHB SSR map was constructed from 264 SSR marker loci, and AFLP markers have a multitude of uses, both are spanned 1199.4 cM, and had a mean density of 4.5 cM per locus. The dominant, multicopy, and often nonspecific in nature RHA280 ϫ RHA801 map was constructed by adding 118 new SSR and, as a whole, unsatisfactory for establishing a geand insertion-deletion (INDEL) marker loci to 459 previously nome-wide framework of DNA markers for anchoring mapped SSR marker loci. The 577-locus map spanned 1423.0 cM and cross referencing genetic linkage maps. Single-copy, and had a mean density of 2.5 cM per locus. The three maps were codominant DNA markers, e.g., SSRs, are preferred for constructed from 1044 DNA marker loci (701 unique SSR and 89 such purposes and, until recently, have been lacking unique RFLP or INDEL marker loci) and supply a dense genomein sunflower. wide framework of sequence-based DNA markers for molecular breeding and genomics research in sunflower.

Background: Earlier comparative maps between the genomes of rice (Oryza sativa L.), barley (Hordeum vulgare L.) and wheat (Triticum aestivum L.) were linkage maps based on cDNA-RFLP markers. The low number of polymorphic RFLP markers has limited the development of dense genetic maps in wheat and the number of available anchor points in comparative maps. Higher density comparative maps using PCR-based anchor markers are necessary to better estimate the conservation of colinearity among cereal genomes. The purposes of this study were to characterize the proportion of transcribed DNA sequences containing simple sequence repeats (SSR or microsatellites) by length and motif for wheat, barley and rice and to determine in-silico rice genome locations for primer sets developed for wheat and barley Expressed Sequence Tags.

Expressed sequence tags (ESTs) are a valuable source of molecular markers. To enhance the resolution of an existing linkage map and to identify putative functional polymorphic gene loci in hexaploid wheat (Triticum aestivum L.), over 260,000 ESTs from 5 different grass species were analyzed and 5418 SSR-containing sequences were identified. Using sequence similarity analysis, 156 cross-species superclusters and 138 singletons were used to develop primer pairs, which were then tested on the genomic DNA of barley (Hordeum vulgare), maize (Zea mays), rice (Oryza sativa), and wheat. Three-hundred sixty-eight primer pairs produced PCR amplicons from at least one species and 227 primer pairs amplified DNA from two or more species. EST-SSR sequences containing dinucleotide motifs were significantly more polymorphic (74%) than those containing trinucleotides (56%), and polymorphism was similar for markers in both coding and 5' untranslated (UTR) regions. Out of 112 EST-SSR markers, 90 identified 149 loci that were integrated into a reference wheat genetic map. These loci were distributed on 19 of the 21 wheat chromosomes and were clustered in the distal chromosomal regions. Multiple-loci were detected by 39% of the primer pairs. Of the 90 mapped ESTs, putative functions for 22 were identified using BLASTX queries. In addition, 80 EST-SSR markers (104 loci) were located to chromosomes using nullisomic-tetrasomic lines. The enhanced map from this study provides a basis for comparative mapping using orthologous and PCR-based markers and for identification of expressed genes possibly affecting important traits in wheat.

Simple sequence repeat (SSR) markers were developed for cultivated sunflower (Helianthus annuus L.) from the DNA sequences of 970 clones isolated from genomic DNA libraries enriched for (CA)n,, (CT)n, (CAA)n, (CATA)n, or (GATA)n. The clones harbored 632 SSRs, of which 259 were unique. SSR markers were developed for 130 unique SSRs by designing and testing primers for 171 unique SSRs. Of the total, 74 SSR markers were polymorphic when screened for length polymorphisms among 16 elite inbred lines. The mean number of alleles per locus was 3.7 for dinucleotide, 3.6 for trinucleotide, and 9.5 for tetranucleotide repeats and the mean polymorphic information content (PIC) scores were 0.53 for dinucleotide, 0.53 for trinucleotide, and 0.83 for tetranucleotide repeats. Cluster analyses uncovered patterns of genetic diversity concordant with patterns produced by RFLP fingerprinting. SSRs were found to be slightly more polymorphic than RFLPs. Several individual SSRs were significantly more polymorphic than RFLP and other DNA markers in sunflower (20% of the polymorphic SSR markers had PIC scores ranging from 0.70 to 0.93). The newly developed SSRs greatly increase the supply of sequence-based DNA markers for DNA fingerprinting, genetic mapping, and molecular breeding in sunflower; however, several hundred additional SSR markers are needed to routinely construct complete genetic maps and saturate the genome.