Genome sequencing and the associated bioinformatics is now a widely accepted research tool for accelerating genetic research and the analysis of genome structure and function of wheat because it leverages similar work from other crops and plants. The International Wheat Genome Sequencing Consortium addresses the challenge of wheat genome structure and function and builds on the research efforts of Professor Bob McIntosh in the genetics of wheat. Currently, expressed sequence tags (ESTs; similar to 500 000 to date) are the largest sequence resource for wheat genome analyses. It is estimated that the gene coverage of the wheat EST collection is similar to 60%, close to that of Arabidopsis, indicating that similar to 40% of wheat genes are not represented in EST collections. The physical map of the D-genome donor species Aegilops tauschii is under construction ( http:// wheat. pw. usda. gov/ PhysicalMapping). The technologies developed in this analysis of the D genome provide a good model for the approach to the entire wheat genome, namely compiling BAC contigs, assigning these BAC contigs to addresses in a high resolution genetic map, filling in gaps to obtain the entire physical length of a chromosome, and then large-scale sequencing.

Molecular markers have provided plant breeders with the molecular tools to select for plants with the required quality and disease traits during plant breeding. The challenge to plant breeders is to implement these molecular markers into modern breeding programs in a cost-effective and efficient way. Essentially, the screening of a plant during marker-assisted selection (MAS) in a wheat breeding program is a four-step process: (1) collection of leaf samples from the plant to be screened in the field plot or glasshouse trial; (2) the extraction of DNA from the leaf sample; (3) the PCR-based marker assay and fragment analysis, and finally (4) the integration of marker results into breeder’s database. The ability to automate each step is a highly desirable aim to enable the screening of thousands of plants required annually by plant breeders in large breeding populations. This presentation will outline the application of high-throughput methodologies such as DNA extraction from leaf samples collected in a 96 well format, use of the Beckman Biomek 2000 robotic workstation to set up the PCR marker assay, and barcoding to track samples from the field plot to the lab bench. Streamlining of this process will considerably reduce the cost and increase the efficiency of marker-assisted breeding for wheat breeders, ultimately leading to the aim of the release of wheat cultivars with superior agronomic characteristics.

Molecular markers have provided plant breeders with the molecular tools to select for plants with the required quality and disease traits during plant breeding. The challenge to plant breeders is to implement these molecular markers into modern breeding programs in a cost-effective and efficient way. Essentially, the screening of a plant during marker-assisted selection (MAS) in a wheat breeding program is a four-step process: (1) collection of leaf samples from the plant to be screened in the field plot or glasshouse trial; (2) the extraction of DNA from the leaf sample; (3) the PCR-based marker assay and fragment analysis, and finally (4) the integration of marker results into breeder’s database. The ability to automate each step is a highly desirable aim to enable the screening of thousands of plants required annually by plant breeders in large breeding populations. This presentation will outline the application of high-throughput methodologies such as DNA extraction from leaf samples collected in a 96 well format, use of the Beckman Biomek 2000 robotic workstation to set up the PCR marker assay, and barcoding to track samples from the field plot to the lab bench. Streamlining of this process will considerably reduce the cost and increase the efficiency of marker-assisted breeding for wheat breeders, ultimately leading to the aim of the release of wheat cultivars with superior agronomic characteristics.

Late maturity α−amylase (LMA) refers to the synthesis of α−amylase activity, which occurs under particular environmental conditions during the later stages of grain ripening in some genotypes in the absence of rain or sprouting. Grain affected by LMA may have a sound appearance, but because of the high α−amylase activity can be unsuitable for a range of end-product applications. Screening for LMA in wheat cultivars is difficult due to the influence of the environment on the expression of this trait. Therefore, the development of a molecular marker to .tag. this trait for marker assisted selection in wheat breeding programs would be a definite advantage. Bulked segregant analysis (BSA) and fluorescent amplified fragment length polymorphism (AFLP) technologies were used to identify loci linked to LMA in a doubled haploid population derived from wheat (Triticum aestivum L. em. Thell) cultivars Cleo-Inia (LMA source) and Janz (non-LMA). Markers linked to LMA were identified in bulks developed from 10 LMA and 10 non-LMA plants, converted to PCR based markers using radioactive AFLP and validated on other cultivars with known LMA phenotype.

Five sets of markers were assessed for their usefulness in breeding, two linked to wheat stem rust gene Sr2, several markers linked to a chromosome segment conferring Yr17/Lr37/Sr38 resistance, two reported markers for the linked genes Lr35 andSr39, one for Lr28, and one linked to flour colour. The gene for Sr2 confers adult plant resistance to stem rust (Puccinia graminis f.sp. tritici) and was originally transferred to bread wheat from the tetraploid emmer (‘Yaroslav’) to the cultivars Hope and H-44. The gene is located on the short arm of chromosome 3B and confers a durable adult plant resistance to stem rust usually expressed only in the field. The chromosome segment carrying the Lr37, Sr38, Yr17 resistance genes is located on 2AS and was originally introduced into wheat through an Aegilops ventricosa Triticum persicum cross, followed by a cross to the cultivar Marne (VPM1). The flour colour quantitative trait locus was originally described in a Yarralinka Schomburg cross and is located on chromosome 7A. The primers as originally developed required optimisation for more routine use in a breeding program.

Two PCR-based assays were examined for tracing the presence of a Thinopyrum chromosome segment (Tc6 or Tc14) conferring barley yellow dwarf virus (BYDV) resistance in wheat breeding lines. The microsatellite gwm37 was used to assay the Thinopyrum chromosome segment or its wheat, Group 7, homoeologous segment, and was effective in characterising breeders material since heterozygous lines could be identified. A new set of primers derived from a Thinopyrum-specific DNA segment (csTiB1) provided a dominant marker that was readily scored by agarose gel electrophoresis. It was also demonstrated that the csTiB1 primers could be used to establish a solid phase PCR assay that avoided the requirement for gel electrophoresis and was amenable to use in a high-throughput, microtitre plate format. Depending on the number of DNA samples to be assayed, both primer pairs appear to have a place in breeding programs.

The absence of expression of the granule-bound starch synthase I (GBSSI) allele from chromosome 4A of wheat is associated with improved starch quality for making Udon noodles. Several PCR-based methods for the analysis of GBSS alleles have been developed for application in wheat. A widely applied approach has involved a simple PCR followed by electrophoretic separation of DNA products on agarose gels. The PCR amplifies one band from each of the loci on chromosomes 4A (Wx-B1), 7A (Wx-A1), and 7D (Wx-D1), and the band from the Wx-B1 locus is diagnostic for the occurrence of the null Wx-B1 allele that is associated with improved starch quality. The reliable detection of the null Wx-B1 allele has been important in identifying wheat breeding lines. Allele-specific PCR has also been used to successfully detect the occurrence of the null Wx-B1 allele. In the present paper the various protocols were evaluated by testing a segregating double haploid population from a cross between Cranbrook and Halberd and the tests gave good agreement in different laboratories. The application of the DNA-based tests applied in wheat breeding programs provides one of the first examples of a molecular marker selection for a grain quality trait being successfully applied in an Australian wheat breeding program.