1a.Objectives (from AD-416):
It may be possible to use gametic incompatibility systems to prevent corn from being pollinated by unwanted pollen. We will test this hypothesis by development of corn varieties containing combinations of gametic incompatibility systems. The objectives of this agreement are to develop functional markers for all known alleles at the Ga1 locus and to design a robust phenotyping strategy that will allow development of the desired varieties.
1b.Approach (from AD-416):
The cooperator is engaged in map-based cloning of the Ga1 locus. Functional markers will be designed based on the sequences of the ga1, Ga1-S and Ga1-M alleles once these sequences are available. A phenotyping strategy containing positive and negative controls for the different gametic incompatibility loci will be developed and tested in our nursery this summer. This strategy involves pollinating plants to be phenotyped by pollen mixtures that have been characterized by pollination of a series of control plants.
Iowa State University (ISU) developed and tested a number of genetic markers that define the general location of the Ga1-s allele in a different genetic background. They switched mapping populations and are now working in the W22 public inbred background for all mapping and phenotyping. Switching genetic backgrounds makes pollinations simpler as all the mapping populations and positive and negative control lines are also in the W22 background so they all flower at about the same time in the nursery. This background also has some better agronomic qualities (e.g., less stalk lodging) that also make it more efficient to work within the nursery. Since ISU changed backgrounds, they needed to test that all the markers were polymorphic (different) in this new background and the markers could be used with a high throughput marker analysis technology (called HRM for High Resolution Melt). Thus, several months were spent optimizing marker design, testing each marker and optimizing parameters for the HRM technology to enable robust and efficient marker analysis. ISU performed a pilot mapping experiment in the greenhouse on a limited size BC1 (backcross) mapping population (ca. 300 plants) using the markers optimized for HRM. Several informative recombinants (i.e., plants with genetic crossovers within this region which help delimit the precise location of the Ga1-s allele) were identified and self-pollinated in the greenhouse. Progeny rows will be phenotyped in the winter nursery. A much larger BC1 mapping population is currently growing in the summer nursery in Ames, Iowa (ca. 3000 plants) and will be assayed for informative recombinant plants with the HRM-optimized markers. Lastly, several Bacterial Artificial Chromosome (BAC) clones containing large pieces of maize genomic DNA that comprise parts of the Ga1-s region have been sequenced. The sequences are being assembled in order to determine if this region of genomic maize DNA with the Ga1-s allele is markedly different than this region from a line (B73) with a normal ga1 allele that is available online from the maize genome sequence project.