Objective 1: Evaluation of the effect of environmental factors in CO frequency in wheat and wild relative species: To assess the effect of nutrient concentration on homologous and homoeologous CO frequency in meiotic metaphase I, we added a modified Hoagland solution to the soil in which wheat and wheat-rye hybrids. In the absence of Ph1, we can increase the CO frequency by altering environmental factors: we show that higher nutrient levels in the soil or lower temperatures increase the level of both homologue and homoeologue COs. In the presence of Ph1, no significant increase in CO number was observed.
Objective 2: Exploitation of the ZIP4 homologue within the wheat Ph1 locus: The wheat Ph1 locus promotes accurate synapsis and CO of homologous chromosomes. Interspecific hybrids between wheat and wild relatives are exploited by breeders to introgress important traits from wild relatives into wheat, although in hybrids between hexaploid wheat and wild relatives, which possess only homoeologues, COs do not take place during meiosis at metaphase I. However, in hybrids between Ph1 deletion mutants and wild relatives, COs do take place. A single Ph1 deletion (ph1b) mutant has been exploited for the last 40 years for this activity. We show here that chemically induced mutant lines, selected for a mutation in TaZIP4-B2 within the Ph1 locus, exhibit high levels of homoeologous crossovers when crossed with wild relatives. Tazip4-B2 mutant lines may be more stable over multiple generations, as multivalents causing accumulation of chromosome translocations are less frequent. Exploitation of such Tazip4-B2 mutants, rather than mutants with whole Ph1 locus deletions, may therefore improve introgression of wild relative chromosome segments into wheat.
Objective 3: Identification of the macronutrient responsible for the increase in the CO number observed in ZIP4 (Ph1) mutant wheat-wild relative hybrids: Wild relatives provide an important source of useful traits in wheat breeding. Wheat and wild relative hybrids have been widely used in breeding programs to introduce such traits into wheat. However, successful introgression is limited by the low frequency of homoeologous CO between wheat and wild relative chromosomes. Hybrids between wheat carrying a 70Mb deletion (ph1b) and wild relatives, have been exploited to increase the level of homoeologous CO, allowing chromosome exchange between their chromosomes. In ph1b-rye hybrids, CO number increases from a mean of 1 CO to 7 COs per cell and up to a mean of 12 COs per cell by treating the plants with Hoagland solution. More recently, it was shown that the major meiotic CO gene ZIP4 (TaZIP4-B2) within the 70Mb deletion, was responsible for the restriction of homoeologous COs in wheat-wild relative hybrids, confirming the ph1b phenotype as a complete Tazip4-B2 deletion mutant (Tazip4-B2 ph1b). We have identified the particular Hoagland solution constituent responsible for the increased chiasma frequency in Tazip4-B2 ph1b mutant-rye hybrids and extended the analysis to Tazip4-B2 TILLING and CRISPR mutant-Ae variabilis hybrids. A significant increase of homoeologous CO frequency was observed in all analysed hybrids, when plants were irrigated with a 1mM Mg2+ solution. These observations suggest a role for Mg2+ supplementation in improving the success of genetic material introgression from wild relatives into wheat.
